Novel Conjugated Quinoline–Indoles Compromise Plasmodium falciparum Mitochondrial Function and Show Promising Antimalarial Activity

Article pubs.acs.org/jmc

Novel Conjugated Quinoline−Indoles Compromise Plasmodium falciparum Mitochondrial Function and Show Promising Antimalarial Activity Silvia C. Teguh,†,∥ Nectarios Klonis,†,∥ Sandra Duffy,# Leonardo Lucantoni,# Vicky M. Avery,# Craig A. Hutton,§,∥ Jonathan B. Baell,*,⊥,▽ and Leann Tilley*,†,‡,▽ †

Department of Biochemistry and Molecular Biology, ‡ARC Centre of Excellence for Coherent X-ray Science, and §School of Chemistry, ∥Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne VIC 3010, Australia ⊥ Medicinal Chemistry, Monash Institute of Pharmaceutical Science, Parkville VIC 3052, Australia # Eskitis Institute for Drug Discovery, Brisbane Innovation Park, Griffith University, Nathan QLD 4111, Australia S Supporting Information *

ABSTRACT: A novel class of antimalarial compounds, based on an indol-3-yl linked to the 2-position of a 4-aminoquinoline moiety, shows promising activity against the malaria parasite, Plasmodium falciparum. Compounds with a quaternary nitrogen on the quinoline show improved activity against the chloroquine-resistant K1 strain. Nonquaternerized 4-aminoquinolines retain significant potency but are relatively less active against the K1 strain. Alkylation of the 4-amino group preferentially improves the activity against the chloroquinesensitive 3D7 strain. The quinoline-indoles show only weak activity as inhibitors of β-hematin formation, and their activities are only weakly antagonized by a hemoglobinase inhibitor. The compounds appear to dissipate mitochondrial potential as an early event in their antimalarial action and therefore may exert their activity by compromising Plasmodium mitochondrial function. Interestingly, we observed a structural relationship between our compounds and the anticancer and anthelminthic compound, pyrvinium pamoate, which has also been proposed to exert its action via compromising mitochondrial function.

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INTRODUCTION Malaria remains one of the world’s greatest global health challenges. It is estimated that 2.2 billion people live in malaria endemic areas and that the more than 500 million cases of malaria each year result in up to 1.2 million deaths, mostly in young children.1,2 Plasmodium falciparum is the causative agent of the most severe form of malaria in humans. Drugs have long been a mainstay in the fight against malaria, but their use has been dogged by the development of resistance. While a number of highly successful drugs have been deployed (including quinoline and sulfa drugs), the rapid development and spread of resistance has rendered most of them ineffective in endemic areas.3 For example until the advent of widespread resistance, the efficacy, long half-life, affordability, and safety profile of chloroquine (3, Figure 1) and other 4-aminoquinolines made this the drug class of choice for combating malaria. 4Aminoquinolines target the pathway for detoxification of hemoglobin breakdown products.4−6 Chloroquine, a weak base, is accumulated in the parasite’s acidic digestive vacuole.7 Here it binds the hematin that is released during hemoglobin degradation and prevents the formation of hemozoin.8−11 Free hematin has redox and detergent-like activities that are thought to lead to rapid parasite killing.12−15 © 2013 American Chemical Society

Unfortunately chloroquine resistance emerged after decades of successful use of this drug and has since reached all malariaendemic regions.16 Chloroquine resistance arises as a consequence of mutations in a transporter in the digestive vacuole membrane (called the P. falciparum chloroquine resistance transporter, PfCRT), with the mutant protein mediating the export of the drug out of the vacuole and hence away from its site of action.17−19 The effectiveness of chloroquine has now declined to the point where it has been officially abandoned in most countries. Nonetheless, newer drugs are too expensive for many and 300−500 million courses of chloroquine are still used each year.20−23 Other antimalarials such as atovaquone target the mitochondrial cytochrome bc1 complex, while antifolate agents inhibit dihydropteroate synthase or dihydrofolate reductase;24,25 however, widespread resistance has reached the level where these drugs have only limited use.26,27 The World Health Organization currently recommends artemisinin-based combination therapies (ACTs) for treating uncomplicated malaria.1 ACTs combine an artemisinin derivative, with another longer-acting drug such as lumefantrine, Received: May 3, 2013 Published: July 9, 2013 6200

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Figure 1. Structures of the main compounds discussed in this study.

Scheme 1. Synthesis of the Target Aminoquinolium−Indole Conjugatesa,b

Reagents and conditions: (a) EtOH, 120 °C, microwave, 4 h; (b) Ac2O, pyridine, reflux, 4−24 h; (c) MeI or BuI, acetonitrile, reflux, 24−48 h; (d) 9a−e, (1) CH3CN or EtOH, piperidine (2 equiv), reflux, overnight; optional additional condition, (2) K2CO3, MeOH, stirring, 5−24 h. bYields and substituents: see Table 1. a

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RESULTS Chemistry. We previously identified an N-acetylated analogue of quinolinium 1a (Scheme 1, Table 1) from a high throughput screen (HTS) as having antimalarial activity.38 Subsequent synthetic studies revealed that the N-acetyl group of the HTS hit could be removed without loss of activity (Teguh, Hutton, Baell, unpublished data). This compound, 1a, represents a better starting point for SAR elaboration as we foresaw the Nacyl group as being potentially reactive. The origin of the acetylated compound was not recorded in prior literature, and no synthetic route was available.38 We found that the synthetic route shown in Scheme 1, adapted from some related synthetic manipulations, served as a suitable basis for most of the desired SAR probes of 1a.39−41 This involves acetylation of the 4-amino group and then ring N-methylation of 4-aminoquinaldine (6) to give 7 then 8, respectively, followed by a one-pot condensation with an aryl aldehyde (9, in the presence of 2 equiv of piperidine) that, in most cases, simultaneously leads to the deacetylation of the resulting conjugate to give final products 1a−h. Most 4aminoquinaldine and aryl aldehyde precursors were obtained commercially. Otherwise, they were synthesized from the corresponding 4-chloroquinaldine.42 The likely mechanism is shown in Scheme 2, with the key step involving attack of a reactive quaternary Schiff’s base (formed through condensation of piperidine with aryl aldehyde) with the carbanion of the acetylated 4-aminoquinaldine (for review, see ref 43). Using ethanol as the solvent for the condensation reaction, as suggested in the literature for related systems, resulted in lower than

amodiaquine, mefloquine, or sulfadoxine−pyrimethamine, with a different mode of action.28 Because current antimalarial control in endemic areas is highly dependent on ACTs, recent reports of decreased clinical efficacy of artemisinin and its derivatives are extremely concerning.29−31 Increased efforts in antimalarial drug discovery have been supported by a variety of initiatives such as those involving the Wellcome Trust, Medicines for Malaria Venture, and the Genomic Institute of the Novartis Research Foundation. Some drug companies such as GlaxoSmithKline have screened their corporate libraries and disclosed the structures of antimalarial compounds. These large-scale studies involved screening hundreds of thousands and, in one study, almost two million compounds.33,32 These studies have yielded molecules that are potent inhibitors of the growth of blood stage P. falciparum, and preclinical and clinical candidates are emerging.34−37 Nevertheless, there remains a critical need for the development of novel effective antimalarial drugs with efficacy against chloroquine-resistant strains. We recently reported the serendipitous discovery of an indole-linked quinolinium with promising activity against P. falciparum.38 We now report our investigation into the structure−activity relationships (SAR) of this compound class. We have determined antimalarial activities against chloroquine-sensitive and chloroquine-resistant strains and investigated the mechanism of action. 6201

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Table 1. Intermediates and Final Compounds and Reaction Yields R1

compd no

8

R4

R5

Y

yield (%)

H Me H CH(CH3)CH2CH2CH2N(Et)2

H H 7-Cl H

a b c

H Me H

H H 7-Cl

a b c d

H Me H H

H H 7-Cl H

a b c d e

1

a b c d e f g h

H H Me H H H H H

methodb

*

a b c d

9

notea *

Me

5

7

R3

H

4

6

R2

H H H H H H 7-Cl H

* 99 69 67 75 84 Me Me Me Bu

Me Me Me Me Me Me Me Bu

82 59 32 60 Me H H H H

H H 6-F H H

C C C [7-N] [2-N]

Me H Me H H H H H

H H H 6-F H H H H

C C C C [7-N] [2-N] C C

* *

Scheme 5

̂ ̂ ̂ ̂ ̂ ̂ * * * * *

58 57 44 20 16 62 73 59 ̂

̂ ̂ ̂ ̂ ̂ ̂ ̂

a a b a a c d c

The meaning of the symbols: * = literature procedure; ̂ = novel compounds; square brackets [ ] = the nitrogen is replacing the aromatic carbon atom. bMethods: a = ethanol used as solvent without second step using K2CO3; b = ethanol with second step; c = acetonitrile solvent without second step; d = acetonitrile solvent with second step a

Scheme 2. Plausible Mechanism for the Condensation Reaction

Scheme 3. The Unsuccessful Attempt to Perform the Condensation without the N-Acetyl Groupa

a

Reagents and conditions: (a) CH3I, CH3CN, reflux, overnight; (b) 9a, piperidine, EtOH or CH3CN, reflux overnight.

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Scheme 4. Attempted Route to Obtain Neutral Quinoline−Indole Vinylic Conjugate (11a) in One Stepa

Reagents and conditions: (a) piperidine (14 mol %), CH3CN, microwave, 158 °C, 4.5 h; (b) (1) ZnCl2, 185 °C 2 h, (2) 1 M HCl(aq), reflux, 0.5 h; (c) DABCO, EtOH, reflux, overnight.

a

Scheme 5. Synthesis of Neutral Conjugated Targets, Including Chloroquine-Hybrid 11c and Reduced Product 15a

Reagents and conditions: (a) Boc2O (2 equiv), THF, NaHMDS, 0−25 °C, stirring, 24 h; (b) EtOH, microwave, 220 °C, 8 h;. (c) 9a and 9e, 14% piperidine, CH3CN, microwave, 158 °C, 4 h; (d) MeOH, 10% Pd/C (cat.), H2(g), stirring, overnight.

a

expected yields (20−40%).39 We attributed this to premature loss of the N-acetyl group before condensation, either through base-assisted solvolysis or through transfer to piperidine (the latter also causing deactivation of piperidine) or a combination of the two. Attempts to replace piperidine with a less nucleophilic base such as triethylamine failed, presumably because it could not form the key quaternary iminium species to allow the reaction to take place. To investigate whether solvolysis was the dominating issue, ethanol was replaced with a solvent which would not attack the N-acetyl group. Acetonitrile was chosen, resulting in consistently improved yields of condensed product (by ∼2fold), although subsequent treatment with potassium carbonate in methanol was required to then remove the N-acetyl group. Relevant target compounds and intermediates and their yields are listed in Table 1. Shortening the synthetic route by obviating the acetylation/ deacetylation steps was attempted using 10 (which was readily generated from 6a as shown), but this yielded no desired product in the condensation step (Scheme 3). This is presumably due to the decrease in the acidity of the methyl proton of the quinolinium species 10 relative to that in its activated counterpart 8a, preventing formation of the carbanion. We also targeted the synthesis of nonquaternary salt forms of these compounds, such as the neutral quinoline−indole vinylic conjugate 11a. We initially attempted condensation using our previously successful conditions on nonactivated precursors to see if we could directly

obtain 11a, but not unexpectedly (given the proposed mechanism), we only obtained traces of the desired product (Scheme 4). The literature describes a variety of methods to construct related systems. For example, condensation has been catalyzed by Lewis acids such as zinc(II) chloride to give styrene derivatives.44 Alternatively, the N-methyl group of the quaternary salt in related compounds has been removed after condensation with a strong base such as DABCO and pyridinium chloride.45,46 However, none of these methods were successful for synthesizing our compounds (Scheme 4). Instead, we investigated alternative ways to activate quinoline precursor 6a. This was achieved through installation of two Boc groups on the 4-amino group, giving 13, and the reaction was conducted using microwave conditions, as shown in Scheme 5.47 Condensation proceeded with concomitant cleavage of the Boc protecting groups to furnish 11a in an acceptable yield of ∼28%. We also prepared the alkyl-linked derivative 15 via reduction of the double bond of 11a. The conjugated indazole 11b was also synthesized, albeit in low yield, using this approach. Finally, we incorporated a chloroquine-like side chain in these compounds via displacement of the labile halogen in 4 with amine 14, giving desired product 11c using the same microwavemediated condensation conditions (Scheme 5). Antimalarial Activity. We examined the antimalarial activity of the novel compounds against chloroquine-sensitive and resistant strains (3D7 and K1, respectively) during a 48 h 6203

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Table 2. Growth Inhibition of Chloroquine-Sensitive (3D7) and Chloroquine-Resistant (K1) P. falciparuma and Selectivity Indices against HEK293 Mammalian Cellsb

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Table 2. continued Determined for 48 h exposure using SYBR Green-I fluorescence method. The ± values, where given, indicate the standard deviations of three individual experiments. bSelectivity index values based on HEK293 and 3D7 P. falciparum viability assays using a 72 h exposure period (see Table S1, Supporting Information, for raw values). n/a: not available. cNo activity at the highest concentration tested. a

and 11b were 2−3-fold less potent than their respective quinolinium counterparts 1b and 1f against the K1 strain but 2−3-fold more active against the 3D7 strain. They also maintain good selectivity against P. falciparum relative to mammalian cells. The indazole ring was common to the two most selective compounds, 1f and 11b, suggesting that this moiety is an important determinant of lower mammalian cytotoxicity. The change from an N-alkylquinolinium to a quinoline ring thus appears to make the compounds more chloroquine-like with respect to their activities toward chloroquine-sensitive and resistant strains, although the RI of the compounds remains superior to that of 3 (2−3 vs 15, respectively). Interestingly, further addition of a chloroquine-like alkylamino side chain to 11a to form 11c produced only a modest (2-fold) increase in activity toward the 3D7 strain (RI ∼3). Because this alkylamino group is expected to enhance accumulation of the compound in the parasite’s digestive vacuole, the site of chloroquine action and location of PfCRT, this suggests that these compounds have a different mode and location of action. We also used the quinoline analogues to examine the influence of the aryl linker on antimalarial activity. Saturation of the alkyl linker (compound 15) was associated with a substantial loss of activity against both strains indicating that the extended conjugation of the two ring systems is important for their activity. Inhibition of β-Hematin Formation. Chloroquine and other 4-aminoquinolines are thought to exert their antimalarial activities by targeting the heme detoxification pathway. We were interested in determining whether the novel compounds target the same pathway. Hemozoin formation in vivo is thought to be facilitated by specific parasite-derived lipids.49−51 In this work, we examined the formation of the synthetic version of hemozoin, βhematin, in the presence of Nonidet P40 detergent, which is a convenient substitute for the lipid catalysts.52 Chloroquine inhibited the formation of β-hematin with an IC50 value of 30 μM (Table 3), which is similar to the IC50 values found using other

incubation period; Table 2). The selectivity of the compounds for P. falciparum (3D7) compared to mammalian (HEK293) cells was determined using 72 h assays, with the selectivity index (SI) defined as the ratio of the IC50 values (Table 2 and Table S1, Supporting Information). Compound 1a inhibits the growth of the 3D7 strain with an IC50 value of 170 nM, in agreement with previous data for the N-acetylated derivative.38 Importantly, this compound shows similar activity against the K1 strain (resistance index, RI, of 1.3 compared with 15 for 3) and reasonable selectivity for P. falciparum (selectivity index, SI, of 18). We first examined the influence of the indole ring and its substituents on antimalarial activity and selectivity. A compound devoid of the indole ring (10) was associated with complete loss of activity against both P. falciparum strains, indicating this is critical to the antimalarial activity of these compounds. A compound without the 2-methyl group in the indole ring (1b) did not affect the activity against the 3D7 strain but resulted in ∼2-fold increase in activity against the K1 strain (RI = 0.6) and a modest improvement in selectivity (SI = 35). Further changes to the indole structure, including replacement of the indole ring with indazole (1f), the 7-aza heterocycle (1e), or the addition of a 6-fluorine substituent (1d), produced relatively small decreases in activity against both parasite strains (typically 4000 1100 ± 190 30 ± 10

The ± values, where given, indicate the range of variability between two individual experiments (the values for each experiment are the average of triplicate determinations).

a

methods of promoting hematin crystallization.53,54 Inhibition of β-hematin at micromolar concentrations in vitro can lead to antimalarial activity at nanomolar levels in vivo because 3 accumulates in the acidic parasite digestive vacuole, resulting in high intracellular concentrations of active compound.55 The test compounds exhibiting the greatest potency with respect to βhematin inhibition were the quinolinium compounds 1a and 1g 6205

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degradation pathway.56 Using a similar assay design, we exposed trophozoite stage parasites to the test compounds in the presence or absence of ALLN for a period of 5 h. The drug and ALLN were then removed and the parasites cultured for a further 43 h before parasite survival was assessed. The degree of ALLN antagonism is defined as the IC50 ratio measured in the presence and absence of ALLN. In control experiments, we found that treatment of parasites with a pulse of ALLN alone at these concentrations did not cause substantive killing of P. falciparum (data not shown).56 Co-treatment of 3 with 0.2 or 5 μM ALLN had a dramatic effect on its efficacy (Table 4), with values for ALLN antagonism of 4 and 20, respectively. This result is consistent with the proposed mechanism of action of 3 requiring the hemoglobin degradation product, hematin.9,57 In contrast, the antifolate antimalarials, WR99210 and methylene blue, showed little or no interaction with ALLN (antagonism 250 °C, using 8a as the quinolinium species and 9a as the aryl aldehyde and ethanol as the solvent. 1H NMR (500 MHz, DMSO-d6) δ 11.85 (sbr, 1H, indole-NH), 8.55 (sbr, 2H, NH2), 8.42 (d, J = 8.1 Hz, 1H, ArH), 8.18 (d, J = 8.8 Hz, 1H, ArH), 8.05−7.98 (m, 2H, ArH), 7.76 (d, J = 15.6 Hz, 1H, HCC), 7.71 (t, J = 7.6 Hz, 1H, ArH), 7.42 (dd, 1H, ArH), 7.27−7.11 (m, 3H), 7.21 (d, J = 15.2 Hz, 1H, CCH), 4.13 (s, 3H, N-CH3), 2.62 (s, 3H, indole-CH3). 13C NMR (400 MHz, DMSOd6) δ 155.6, 155.1, 142.6, 140.0, 136.2, 134.9, 133.9, 125.5, 125.4, 123.8, 122.1, 121.0, 119.7, 118.5, 116.7, 112.0, 111.6, 109.5, 99.2, 37.1 (NCH3), 12.0 (indole-CH3). IR νmax (cm−1) 3200, 1739, 1631, 1597, 1579, 1561, 1530, 1432, 1226, 738. MS (ESI+) m/z 314.2 (M)+. HRMSa (ESI +) C21H20N3+ requires 314.1657, found 314.1651. Elemental analyses: Found, C, 53.9; H, 4.9; N, 8.5; I, 24.1. C21H20N3I requires C, 57.2; H, 4.6; N, 9.5; I, 28.8%. ( E)-4-Amino-1-methy l-2 -(2-(3a H-indol-3-yl)viny l)quinolinium Iodide (1b). According to general procedure A, the title compound was obtained as brick-orange powder (57% yield), mp > 250 °C, using 8a as the quinolinium species and 9b as the aryl aldehyde and ethanol as the solvent. 1H NMR (300 MHz, DMSO-d6) δ 8.43 (d, J = 8.7 Hz, 1H, ArH), 8.19 (d, J = 8.8 Hz, 1H, ArH), 8.09−7.99 (m, 2H, ArH), 7.79 (d, J = 15.7 Hz, 1H, HCC), 7.72 (t, J = 7.2 Hz, 1H, ArH), 7.52 (dd, J = 7.0 Hz, 1H, ArH), 7.36 (d, J = 15.7 Hz, 1H, CCH), 7.27−7.23 (m, 3H, ArH), 7.14 (s, 1H, ArH), 4.14 (s, 3H, N-CH3). 13C NMR (300 MHz, DMSO-d6) δ 156.0, 155.0, 140.1, 137.3, 135.7, 134.2, 131.3, 125.8, 124.9, 123.9, 122.8, 121.1, 120.0, 118.7, 116.7, 113.7, 113.1, 112.5, 6210

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solvent. 1H NMR (300 MHz, DMSO-d6) δ 8.40 (d, J = 8.9 Hz, 1H, ArH), 8.18 (s, 1H, ArH), 8.05−8.00 (m, 2H, ArH), 7.78 (d, J = 15.6 Hz, 1H, HCC), 7.70 (d, J = 8.1, 1H, ArH), 7.52−7.50 (m, 1H, ArH), 7.23−7.17 (m, 3H, ArH, CCH), 7.07 (s, 1H, ArH), 4.04 (s, 3H, NCH3). 13C NMR (300 MHz, DMSO-d6) δ 155.6, 155.0, 141.0, 138.7, 138.1, 135.9, 132.5, 126.0, 125.6, 125.1, 122.5, 120.9, 119.8, 118.0, 115.7, 113.2, 112.8, 112.4, 99.7, 37.1 (N-CH3). LC-MSc (ESI+) rt 3.4, m/z 334.2 (M)+. HRMSb (ESI+) C20H17ClN3+ requires 334.1107, found 334.1109. (E)-4-Amino-1-butyl-2-(2-(3aH-indol-3-yl)vinyl)quinolinium Iodide (1h). According to general procedure A, the title compound was obtained as light-orange powder (59% yield), using 8d as the quinolinium species and 9b as the aryl aldehyde and acetonitrile as the solvent. 1H NMR (300 MHz, DMSO-d6) δ 8.63 (sbr, 2H, NH2), 8.44 (d, J = 8.7 Hz, 1H, ArH), 8.20 (d, J = 8.9 Hz, 1H, ArH), 8.03−7.98 (m, 2H, ArH), 7.77 (d, J = 16.5 Hz, 1H, HCC), 7.71 (t, J = 7.5 Hz, 1H, ArH), 7.53 (d, J = 7.5 Hz, 1H, ArH), 7.34−7.22 (m, 3H, ArH, CCH), 7.14 (s, 1H, ArH), 4.64 (t, J = 7.7 Hz, 2H, N-CH2CH2CH2CH3), 1.90− 1.78 (m, 2H, N-CH 2 CH 2 CH 2 CH 3 ), 1.54−1.45 (m, 2H, NCH2CH2CH2CH3), 0.97 (t, J = 7.2 Hz, 3H, N-CH2CH2CH2CH3). 13 C NMR (300 MHz, DMSO-d6) δ 156.1, 154.3, 139.2, 137.5, 136.2, 134.4, 131.5, 125.9, 124.9, 124.3, 122.9, 121.1, 119.7, 118.5, 117.0, 113.1, 113.0, 112.7, 99.7, 48.0 (N-CH2), 30.3, 19.3, 13.7 (CH3). LC-MSa (ESI +) rt 6.0, m/z 342.2 (M)+. HRMSb (ESI+) C23H24N3+ requires 342.1973, found 342.1978. Synthesis of N,2-Dimethylquinolin-4-amine (6b).42 4-Chloroquinaldine (4, 0.50 g, 2.8 mmol) and 33% methylamine in ethanol (4.0 mL, 40 mmol) were heated at 120 °C for 7 h in a sealed tube using a microwave reactor. The HCl-salt form of the product was separated by vacuum filtration. It was then treated with excess 5 M KOH(aq), and the pure product was refiltered from the aqueous solution and dried. The product was isolated as yellow−white needles in quantitative yield. 1H NMR (300 MHz, (CD3)2CO) δ 7.99 (dd, J = 8.4 Hz 1H, ArH), 7.76 (dd, J = 8.4 Hz, 1H, ArH), 7.54 (t, J = 8.4 Hz, 1H, ArH), 7.31 (t, J = 8.4 Hz, 1H, ArH), 6.44 (s, 1H, 4-CH3NH), 6.36 (s, 1H, ArH), 2.99 (dbr, 3H, 4CH3NH), 2.50 (s, 3H, CH3). LC-MSa (ESI+) rt 2.0−2.2, m/z 173.2 (M + H)+. Synthesis of N-(5-(Diethylamino)pentan-2-yl)-2-methylquinolin-4-amine (6d).94 4-Chloroquinaldine (4, 1.0 g, 5.6 mmol) and 2amino-5-diethylaminopentane (14, 1.8 g, 5.6 mmol) were dissolved in ethanol (0.50 mL) and heated at 220 °C for 8 h in the microwave. The crude oil was then treated with 5 M KOH, extracted with ethyl acetate, and purified by column chromatography with 10% MeOH in DCM (v/ v) solvent system. The product was isolated as red−brown oil (1.2 g, 69% yield). 1H NMR (300 MHz, CD3OD) δ 11.25 (sbr, 1H, NH), 7.88−7.77 (m, 2H, ArH), 7.61 (t, J = 7.8 Hz, 1H, ArH), 6.83 (s, 1H, ArH), 4.19−4.13 (m, 1H, NHCH(CH3)(CH2)3NEt), 3.32−3.14 (m, 6H, NHCH(CH3)(CH2)3NEt), 2.69 (s, 3H, CH3), 2.01−1.77 (m, 4H, N(CH2CH3)2), 1.41 (d, J = 6.5 Hz, 3H, NHCH(CH3)(CH2)3NEt), 1.28 (t, J = 9.0 Hz, 6H, N(CH2CH3)2). 13C NMR (400 MHz, CDCl3) δ 159.12, 149.12, 148.04, 128.87, 128.59, 123.51, 119.45, 117.38, 98.88, 52.48, 48.00, 46.66 (2C, N(CH2CH3)2), 34.46, 25.45, 23.67, 20.15, 11.30 (2C, N(CH2CH3)2). LC-MSb (ESI+) rt 6.6, m/z 300.1 (M + H)+. N-(2-Methylquinolin-4-yl)acetamide (7a). The title compound was obtained via general procedure B using 4-aminoquinaldine (6a) as light-brown needles (67% yield) mp 161−163 °C (lit. ref mp 163−165 °C).40 1H NMR (500 MHz, CDCl3) δ 8.16 (sbr, 1H, NH), 8.03 (d, J = 7.4 Hz, 1H, ArH), 7.89 (s, 1H, ArH), 7.76 (d, J = 8.3 Hz, 1H, ArH), 7.68 (t, J = 7.7 Hz, 1H, ArH), 7.48 (m, 1H, ArH), 2.72 (s, 3H, 2-CH3), 2.34 (s, 3H, C(O)CH3). MS (ESI+) m/z 201.1 (M + H)+. N-Methyl-N-(2-methylquinolin-4-yl)acetamide (7b). The title compound was obtained via general procedure B using N,2dimethylquinolin-4-amine (6b) as brown powder (75% yield). 1H NMR (300 MHz, CDCl3) δ 7.92 (d, J = 8.3 Hz, 1H, ArH), 7.63−7.54 (m, 2H, ArH), 7.40 (d, J = 7.9 Hz, 1H, ArH), 7.06 (s, 1H, ArH), 3.18 (s, 3H, N(Ac)CH3), 2.61 (s, 3H, 2-CH3), 1.63 (s, 3H, C(O)CH3). 13C NMR (300 MHz, CDCl3) δ 169.5 (CO), 159.5, 149.0, 148.5, 129.8, 128.9, 126.5, 123.0, 121.5, 120.1, 36.3 (N(Ac)CH3), 24.8, 21.5. LC-MSa (ESI+) rt 2.2, m/z 215.4 (M + H)+. HRMSb (ESI+) C13H14N2O requires 214.1116, found 215.1189 (M + H)+.

N-(7-Chloro-2-methylquinolin-4-yl)acetamide (7c). The title compound was obtained via general procedure B using 7-chloro-4amino-2-methylquinoline (6c) as pink−brown powder (84% yield). 1H NMR (300 MHz, (CD3)2CO) δ 9.48 (sbr, 1H, NH), 8.25−8.20 (m, 2H, ArH), 7.91 (s, 1H, ArH), 7.48 (dd, J = 9.0 Hz, 2.4 Hz, 1H, ArH), 2.63 (s, 3H, 2-CH3), 2.30 (s, 3H, C(O)CH3). 13C NMR (300 MHz, CDCl3) δ 169.6 (CO), 161.0, 147.7, 141.8, 136.1, 127.4, 126.6, 121.6, 117.1, 112.3, 25.1, 25.0. LC-MSc (ESI+) rt 2.8, m/z 235.0 (M + H)+. HRMSb (ESI+) C12H11ClN2O requires 234.0564, found 235.0638 (M + H)+. 4-Acetamido-N,2-dimethylquinolinium Iodide (8a). According to general procedure C, the title compound was obtained using quinoline 7a and methyl iodide, being isolated as brown−white powder, (82% yield) mp >250 °C. 1H NMR (400 MHz, DMSO-d6) δ 10.91 (sbr, 1H, NH), 8.77 (d, J = 8.2 Hz, 1H, ArH), 8.63 (s, 1H, ArH), 8.49 (d, J = 8.9 Hz, 1H, ArH), 8.19 (t, J = 7.6 Hz, 1H, ArH), 7.97 (t, J = 7.7 Hz, 1H, ArH), 4.29 (s, 3H, N-CH3), 2.98 (s, 3H, 2-CH3), 2.41 (s, 3H, C(O)CH3). 13C NMR (400 MHz, DMSO-d6) δ 171.2 (CO), 160.2, 148.4, 139.7, 134.6, 127.7, 124.1, 119.0, 118.9, 111.7, 38.8, 25.0, 23.5. IR νmax (cm−1) 3198, 3093, 1718, 1624, 1604, 1580, 1537, 1506, 1344, 1210, 781, 764. MS (ESI+) m/z 215.1 (M)+. HRMS (ESI+) C13H15N2O+ requires 215.1184, found 215.1180. 1,2-Dimethyl-4-(N-methylacetamido)quinolinium Iodide (8b). According to general procedure C, the title compound was obtained using quinoline 7b and methyl iodide, being isolated as brown−yellow powder (59% yield). 1H NMR (300 MHz, CDCl3) δ 8.44 (d, J = 8.8 Hz, 1H, ArH), 8.18 (td, J = 8.8 Hz, 1.5 Hz, 1H, ArH), 8.09 (d, J = 7.1 Hz, 1H, ArH), 8.00 (s, 1H, ArH), 7.91 (t, J = 7.1 Hz, 1H, ArH), 4.61 (s, 3H, N-CH3), 3.56 (s, 3H), 3.34 (s, 3H), 2.18 (s, 3H, C(O)CH3). 13C NMR (300 MHz, DMSO-d6) δ 170.1 (CO), 162.1, 155.9, 140.7, 135.1, 129.3, 125.7, 124.9, 123.9, 119.7, 39.7, 38.0, 23.2, 22.4. LC-MSa (ESI+) rt 0.8, m/z 229.2 (M)+. HRMSb (ESI+) C14H17N2O+ requires 229.1348, found 229.1342. 4-Acetamido-7-chloro-N,2-dimethylquinolinium Iodide (8c). According to general procedure C, the title compound was obtained using quinoline 7c and methyl iodide, being isolated as white−brown powder (32% yield). 1H NMR (300 MHz, DMSO-d6) δ 10.98 (sbr, 1H, NH), 8.80 (d, J = 8.5 Hz, 1H, ArH), 8.63−8.62 (m, 2H, ArH), 8.08 (d, J = 8.5 Hz, 1H, ArH), 4.26 (s, 3H, N−CH3), 2.96 (s, 3H, 2-CH3), 2.39 (s, 3H, C(O)CH3). 13C NMR (300 MHz, DMSO-d6) δ 171.1 (CO), 161.2, 148.4, 140.7, 139.8, 128.2, 126.2, 118.9, 117.7, 112.0, 39.1 (NCH3), 24.9, 23.5. LC-MSc (ESI+) rt 2.7, m/z 249.2 (M)+. HRMSb (ESI +) C13H14ClN2O+ requires 249.0801, found 249.0795. 4-Acetamido-N-butyl-2-methylquinolinium Iodide (8d). According to general procedure C, the title compound was obtained using quinoline 7a and butyl iodide, being isolated as brown−gray powder (60% yield). 1H NMR (300 MHz, CDCl3) δ 10.95 (sbr, 1H, NH), 9.54 (d, J = 8.5 Hz, 1H, ArH), 8.73 (s, 1H, ArH), 8.07−8.01 (m, 2H, ArH), 7.74 (t, J = 7.8 Hz, 1H, ArH), 4.74 (t, J = 8.0 Hz, 2H, NCH2CH2CH2CH3), 3.00 (s, 3H, 2-CH3), 2.75 (s, 3H, C(O)CH3), 1.94−1.86 (m, 2H, NCH 2 CH 2 CH 2 CH 3 ), 1.69−1.59 (m, 2H, NCH2CH2CH2CH3), 1.07 (t, J = 7.2 Hz, 3H, NCH2CH2CH2CH3). 13 C NMR (300 MHz, DMSO-d6) δ 171.2 (CO), 159.6, 148.6, 139.0, 134.8, 127.8, 124.4, 119.3, 119.2, 112.1, 50.1, 30.0, 24.9, 22.8, 19.2, 13.5. LC-MSa (ESI+) rt 4.5, m/z 257.3 (M)+. HRMSb (ESI+) C16H21N2O+ requires 257.1655, found 257.1652. 4-Amino-N,2-dimethylquinolinium Iodide (10). According to general procedure C, the title compound was obtained using quinoline 6a and methyl iodide, being isolated as purple−brown powder (87% yield). 1H NMR (300 MHz, CDCl3) δ 8.76 (sbr, 1H, NH2), 8.44 (d, J = 8.1 Hz, 1H, ArH), 8.17 (d, J = 9.0 Hz, 1H, ArH), 8.02 (t, J = 7.5 Hz, 1H, ArH), 7.73 (t, J = 7.5 Hz, 1H, ArH), 6.73 (s, 1H, ArH), 3.98 (s, 3H, NCH3), 2.72 (s, 3H, 2-CH3). LC-MSc (ESI+) rt 2.6, m/z 173.2 (M)+. 2-((E)-2-(1H-Indol-3-yl)vinyl)quinolin-4-amine (11a). According to general procedure D, using the diBoc-quinaldine (13) and 9b as the aldehyde, the title compound was isolated as orange−yellow powder (28% yield). 1H NMR (300 MHz, CD3OD) δ 8.11 (d, J = 8.1 Hz, 1H, ArH), 7.99−7.92 (m, 2H, ArH, HCC), 7.77−7.76 (m, 2H, ArH), 7.68 (s, 1H, ArH), 7.52−7.39 (m, 2H, ArH), 7.22−7.19 (m, 1H, indole-H), 7.05 (d, J = 16.1 Hz, 1H, CCH), 6.93 (s, 1H, ArH). 13C NMR (300 MHz, CD3OD) δ 157.5, 154.5, 142.2, 139.2, 134.6, 133.7, 131.2, 126.4, 6211

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126.3, 123.9, 123.7, 122.1, 122.0, 121.0, 117.6, 116.2, 114.8, 113.2, 98.3. LC-MSa (ESI+) rt 5.1−5.2, m/z 286.2 (M + H)+. HRMSb (ESI+) C19H15N3 requires 285.1268, found 286.1340 (M + H)+. 2-((E)-2-(1H-Indazol-3-yl)vinyl)quinolin-4-amine (11b). According to general procedure D, using the diBoc-quinaldine (13) and 9e as the aldehyde, the title compound was isolated as yellow powder (∼5% yield). Note that a precipitate was not formed by trituration with dichloromethane according to the general procedure. Therefore the product was isolated by treating the mixture with 33% HCl(aq) and further treating it with K2CO3(aq) (1 equiv) until pH ∼10, after which a precipitate was formed. 1H NMR (400 MHz, MeOD) δ 8.19 (d, J = 8.3 Hz, 1H, ArH), 8.04 (dd, J = 8.4, 0.9 Hz, 1H, ArH), 7.87 (d, J = 8.0 Hz, 1H, ArH), 7.83 (d, J = 16.8 Hz, 1H, HCC), 7.69−7.60 (m, 2H, ArH, CCH), 7.58 (d, J = 8.4 Hz, 1H, ArH), 7.49−7.38 (m, 2H, ArH), 7.29 (td, J = 7.9, 0.9 Hz, 1H, ArH), 7.08 (s, 1H, ArH). 13C NMR (400 MHz, DMSO) δ 157.63, 149.30, 141.43, 140.60, 138.73, 133.94, 131.21, 126.78, 125.98, 123.53, 122.06, 120.98, 120.55, 120.02, 119.75, 115.62, 111.04, 99.00. LC-MSa (ESI+) rt 5.0, m/z 287.2 (M + H)+. HRMSb (ESI +) C18H14N4 requires 286.1224, found 287.1298 (M + H)+. 2-((E)-2-(1H-Indol-3-yl)vinyl)-N-(5-(diethylamino)pentan-2yl)quinolin-4-amine (11c). According to general procedure D, using alkylaminoquinaldine (6d) and 9b as the aldehyde, the title compound was isolated as orange solid (20% yield). 1H NMR (300 MHz, CD3OD) δ 8.40 (d, J = 8.4 Hz, 1H, ArH), 8.27 (d, J = 16.2 Hz, 1H, HCC), 8.07 (dd, 1H, indole-H), 7.86−7.76 (m, 3H, ArH), 7.54 (t, J = 7.7 Hz, 1H, ArH), 7.42−7.39 (m, 1H, indole-H), 7.22−7.14 (m, 3H, ArH, CCH), 7.08 (s, 1H, ArH), 4.34−4.30 (m, 1H, NHCH(CH3)(CH2)3NEt), 3.25−3.03 (m, 6H, NHCH(CH3)(CH2)3NEt), 1.99−1.80 (m, 4H, N(CH2CH3)2), 1.47 (d, J = 6.4 Hz, 3H, NHCH(CH3)(CH2)3NEt), 1.30 (t, J = 7.3 Hz, 6H, N(CH2CH3)2). 13C NMR (300 MHz, CD3OD) δ155.6, 154.0, 139.6, 139.3, 137.2, 134.3, 132.7, 127.1, 126.3, 124.1, 123.7, 122.4, 121.3, 120.3, 117.7, 114.9, 114.0, 113.3, 94.8, 53.0, 50.4, 48.5 (2C), 33.8, 22.2, 20.3, 9.2 (2C). LC-MSb (ESI+) rt 4.1, m/z 427.2 (M + H)+. HRMSb (ESI+) C28H34N4 requires 426.2796, found 427.2863 (M + H)+. Synthesis of 2-Methyl-N,N-di-tert-butoxycarbonylquinolin4-amine (13).95 4-Aminoquinaldine (6a, 1.0 g, 6.3 mmol) was stirred with sodium bis(trimethylsilyl)amide (12 mL of 1 M solution in THF, 12 mmol) in dry THF (30 mL) at 0 °C for the first 0.5 h, then di-t-butyl dicarbonate (2.8 g, 13 mmol) was added and the mixture stirred for a further 24 h. Upon reaction completion, the mixture was treated with 25% NH4Cl(aq), extracted with ethyl acetate, and dried. The crude oil was purified by column chromatography with 15% methanol in DCM solvent system to afford pure product as a yellow oil (40% yield). 1H NMR (300 MHz, CDCl3) δ 8.09 (d, J = 8.4 Hz, 1H, ArH), 7.84 (d, J = 8.1 Hz, 1H, ArH), 7.69 (t, J = 8.4 Hz, 1H, ArH), 7.53 (t, J = 7.9 Hz, 1H, ArH), 7.22 (s, 1H, ArH), 2.77 (s, 1H, 2-CH3), 1.37 (s, 18H, NCOO(C(CH3)3)2. 13C NMR (300 MHz, CDCl3) δ 159.2 (2C, C O), 150.8, 148.8, 144.4, 129.6, 128.8, 126.3, 124.1, 121.7, 121.0, 83.3 (2C, NCOO(C(CH3)3)2), 27.6 (6C, NCOO(C(CH3)3)2), 25.2 (2CH3). LC-MSa (ESI+) rt 6.6, m/z 359.2 (M + H)+. HRMSb (ESI+) C20H26N2O4 requires 358.1895, found 359.1968 (M + H)+. Synthesis of 2-(2-(1H-Indol-3-yl)ethyl)quinolin-4-amine (15) by Hydrogenation.96 11a (20 mg, 0.13 mmol) was dissolved in methanol (3.0 mL), 10% palladium on charcoal (10 mol %) was added, and the mixture was stirred with constant H2(g) flow for 24−30 h (followed by TLC). The palladium on charcoal was then removed by filtration, and the filtrate was dried in vacuo and purified by HPLC. The product was obtained as light-orange powder (53% yield). 1H NMR (300 MHz, CD3OD) δ 8.22 (d, J = 7.8 Hz, 1H, ArH), 7.86 (t, J = 7.5 Hz, 1H, ArH), 7.76 (d, J = 8.1 Hz, 1H, ArH), 7.60 (t, J = 7.8 Hz, 1H, ArH), 7.48 (d, J = 7.5 Hz, 1H, ArH), 7.32 (t, J = 8.1 Hz, 1H, ArH), 7.06 (d, J = 7.5 Hz, 1H, ArH), 7.01 (s, 1H, ArH), 7.60 (t, J = 7.8 Hz, 1H, ArH), 6.58 (s, 1H, ArH), 3.30−3.21 (m, 4H, CH2-CH2). 13C NMR (300 MHz, CD3OD) δ 159.3, 158.7, 141.2, 138.2, 134.5, 128.4, 127.1, 124.1, 123.4, 122.4, 121.2, 119.7, 119.0, 117.0, 113.9, 112.4, 103.0, 36.7, 26.1. LC-MSa (ESI+) rt 5.0, m/z 288.3 (M + H)+. HRMSb (ESI+) C19H17N3 requires 287.1430, found 288.1504 (M + H)+.

Article

ASSOCIATED CONTENT

S Supporting Information *

Additional information includes raw data for selectivity index determination, pKa determination of 11a, concentration− response curves, and mitochondria potential staining figures. This material is available free of charge via the Internet at http:// pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*For L.T.: phone, +61-3-8344-2227; fax, 61-3-9348-1421; Email, [email protected]. For J.B.B.: phone, +61-3-99039044; E-mail, [email protected]. Author Contributions ▽

J.B.B. and L.T. contributed equally. The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS We acknowledge support from the Australian Research Council and the Australian National Health and Medical Research Council. We thank Paul McMillan, Shannon Kenny, Martin Ji, Maria Crespo and James Pham, University of Melbourne, Jason Dang, Monash Institute of Pharmaceutical Sciences, and Brad Sleebs, Danuta Buczek and Wilco Kersten, Walter + Eliza Hall Institute, for technical support and helpful advice.

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ABBREVIATIONS USED

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REFERENCES

ACT, artemisinin combination therapies; SAR, structure− activity relationship; ALLN, N-acetyl-L-leucyl-L-leucyl-L-norleucinal; PfCRT, P. falciparum chloroquine resistance transporter; RI, resistance index; SI, selectivity index

(1) World Malaria Report 2012; World Health Organisation: Geneva, 2012. (2) Murray, C. J. L.; Rosenfeld, L. C.; Lim, S. S.; Andrews, K. G.; Foreman, K. J.; Haring, D.; Fullman, N.; Naghavi, M.; Lozano, R.; Lopez, A. D. Global malaria mortality between 1980 and 2010: a systematic analysis. Lancet 2012, 379, 413−431. (3) Petersen, I.; Eastman, R.; Lanzer, M. Drug-resistant malaria: molecular mechanisms and implications for public health. FEBS Lett. 2011, 585, 1551−1562. (4) Egan, T. J.; Ross, D. C.; Adams, P. A. Quinoline anti-malarial drugs inhibit spontaneous formation of beta-haematin (malaria pigment). FEBS Lett. 1994, 352, 54−57. (5) Slater, A. F.; Swiggard, W. J.; Orton, B. R.; Flitter, W. D.; Goldberg, D. E.; Cerami, A.; Henderson, G. B. An iron-carboxylate bond links the heme units of malaria pigment. Proc. Natl. Acad. Sci. U. S. A. 1991, 88, 325−329. (6) Bray, P. G.; Ward, S. A.; O’Neill, P. M. Quinolines and artemisinin: chemistry, biology and history. Curr. Top. Microbiol. Immunol. 2005, 295, 3−38. (7) Foley, M.; Tilley, L. Quinoline antimalarials: mechanisms of action and resistance and prospects for new agents. Pharmacol. Ther. 1998, 79, 55−87. (8) Bray, P. G.; Janneh, O.; Raynes, K. J.; Mungthin, M.; Ginsburg, H.; Ward, S. A. Cellular uptake of chloroquine is dependent on binding to ferriprotoporphyrin IX and is independent of NHE activity in Plasmodium falciparum. J. Cell Biol. 1999, 145, 363−376. 6212

dx.doi.org/10.1021/jm400656s | J. Med. Chem. 2013, 56, 6200−6215

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(9) Dorn, A.; Stoffel, R.; Matile, H.; Bubendorf, A.; Ridley, R. G. Malarial haemozoin/beta-haematin supports haem polymerization in the absence of protein. Nature 1995, 374, 269−271. (10) Klonis, N.; Dilanian, R.; Hanssen, E.; Darmanin, C.; Streltsov, V.; Deed, S.; Quiney, H.; Tilley, L. Hematin−hematin self-association states involved in the formation and reactivity of the malaria parasite pigment, hemozoin. Biochemistry (Moscow) 2010, 49, 6804−6811. (11) Pagola, S.; Stephens, P. W.; Bohle, D. S.; Kosar, A. D.; Madsen, S. K. The structure of malaria pigment β-haematin. Nature 2000, 404, 307−310. (12) Tilley, L.; Davis, T.; P., B. Prospects for the treatment of drugresistant malaria parasites. Future Microbiol. 2006, 1, 127−141. (13) Becker, K.; Tilley, L.; Vennerstrom, J. L.; Roberts, D.; Rogerson, S.; Ginsburg, H. Oxidative stress in malaria parasite-infected erythrocytes: host−parasite interactions. Int. J. Parasitol. 2004, 34, 163−189. (14) Balla, J.; Vercellotti, G. M.; Jeney, V.; Yachie, A.; Varga, Z.; Jacob, H. S.; Eaton, J. W.; Balla, G. Heme, heme oxygenase, and ferritin: how the vascular endothelium survives (and dies) in an iron-rich environment. Antioxid. Redox Signaling 2007, 9, 2119−2137. (15) Kumar, S.; Bandyopadhyay, U. Free heme toxicity and its detoxification systems in human. Toxicol. Lett. 2005, 157, 175−188. (16) Ecker, A.; Lehane, A. M.; Clain, J.; Fidock, D. A. PfCRT and its role in antimalarial drug resistance. Trends Parasitol. 2012, 28, 504−514. (17) Fidock, D. A.; Nomura, T.; Talley, A. K.; Cooper, R. A.; Dzekunov, S. M.; Ferdig, M. T.; Ursos, L. M.; Sidhu, A. B.; Naude, B.; Deitsch, K. W.; Su, X. Z.; Wootton, J. C.; Roepe, P. D.; Wellems, T. E. Mutations in the P. falciparum digestive vacuole transmembrane protein PfCRT and evidence for their role in chloroquine resistance. Mol. Cell 2000, 6, 861−871. (18) Wootton, J. C.; Feng, X.; Ferdig, M. T.; Cooper, R. A.; Mu, J.; Baruch, D. I.; Magill, A. J.; Su, X. Z. Genetic diversity and chloroquine selective sweeps in Plasmodium falciparum. Nature 2002, 418, 320−323. (19) Martin, R. E.; Marchetti, R. V.; Cowan, A. I.; Howitt, S. M.; Broer, S.; Kirk, K. Chloroquine transport via the malaria parasite’s chloroquine resistance transporter. Science 2009, 325, 1680−1682. (20) Arrow, K. J.; Panosian, C. B.; Gelbrand, H. Committee on the Economics of Antimalarial Drugs Board on Global Health. Saving Lives, Buying Time: Economics of Malaria Drugs in an Age of Resistance; The National Academies Press: Washington, DC, 2004; pp 1−347. (21) Ginsburg, H. Should chloroquine be laid to rest? Acta Trop. 2005, 96, 16−23. (22) Maitland, K.; Makanga, M.; Williams, T. N. Falciparum malaria: current therapeutic challenges. Curr. Opin. Infect. Dis. 2004, 17, 405− 412. (23) Ridley, R.; Toure, Y. Winning the drugs war. Nature 2004, 430, 942−943. (24) Barton, V.; Fisher, N.; Biagini, G. A.; Ward, S. A.; O’Neill, P. M. Inhibiting Plasmodium cytochrome bc1: a complex issue. Curr. Opin. Chem. Biol. 2010, 14, 440−446. (25) Hyde, J. E. Targeting purine and pyrimidine metabolism in human apicomplexan parasites. Curr. Drug Targets 2007, 8, 31−47. (26) Nzila, A. The past, present and future of antifolates in the treatment of Plasmodium falciparum infection. J. Antimicrob. Chemother. 2006, 57, 1043−1054. (27) Muller, I. B.; Hyde, J. E. Antimalarial drugs: modes of action and mechanisms of parasite resistance. Future Microbiol. 2010, 5, 1857− 1873. (28) Enserink, M. Malaria’s drug miracle in danger. Science 2010, 328, 844−846. (29) Dondorp, A. M.; Yeung, S.; White, L.; Nguon, C.; Day, N. P.; Socheat, D.; von Seidlein, L. Artemisinin resistance: current status and scenarios for containment. Nature Rev. Microbiol. 2010, 8, 272−280. (30) Phyo, A. P.; Nkhoma, S.; Stepniewska, K.; Ashley, E. A.; Nair, S.; McGready, R.; ler Moo, C.; Al-Saai, S.; Dondorp, A. M.; Lwin, K. M.; Singhasivanon, P.; Day, N. P.; White, N. J.; Anderson, T. J.; Nosten, F. Emergence of artemisinin-resistant malaria on the western border of Thailand: a longitudinal study. Lancet 2012, 379, 1960−1966.

(31) Stepniewska, K.; Ashley, E.; Lee, S. J.; Anstey, N.; Barnes, K. I.; Binh, T. Q.; D’Alessandro, U.; Day, N. P.; de Vries, P. J.; Dorsey, G.; Guthmann, J. P.; Mayxay, M.; Newton, P. N.; Olliaro, P.; Osorio, L.; Price, R. N.; Rowland, M.; Smithuis, F.; Taylor, W. R.; Nosten, F.; White, N. J. In vivo parasitological measures of artemisinin susceptibility. J. Infect. Dis. 2010, 201, 570−579. (32) 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-Diaz, M. B.; Martinez, M. S.; Wilson, E. B.; Tripathi, A. K.; Gut, J.; Sharlow, E. R.; Bathurst, I.; El Mazouni, F.; 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.; Van Voorhis, W. C.; Avery, V. M.; Guy, R. K. Chemical genetics of Plasmodium falciparum. Nature 2010, 465, 311− 315. (33) Gamo, F. J.; Sanz, L. M.; Vidal, J.; de Cozar, C.; Alvarez, E.; Lavandera, J. L.; Vanderwall, D. E.; Green, D. V.; 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. (34) Rottmann, M.; McNamara, C.; Yeung, B. K.; Lee, M. C.; Zou, B.; Russell, B.; Seitz, P.; Plouffe, D. M.; Dharia, N. V.; Tan, J.; Cohen, S. B.; Spencer, K. R.; Gonzalez-Paez, G. E.; Lakshminarayana, S. B.; Goh, A.; Suwanarusk, R.; Jegla, T.; Schmitt, E. K.; Beck, H. P.; Brun, R.; Nosten, F.; Renia, L.; Dartois, V.; Keller, T. H.; Fidock, D. A.; Winzeler, E. A.; Diagana, T. T. Spiroindolones, a potent compound class for the treatment of malaria. Science 2010, 329, 1175−1180. (35) Younis, Y.; Douelle, F.; Feng, T. S.; Gonzalez Cabrera, D.; Le Manach, C.; Nchinda, A. T.; Duffy, S.; White, K. L.; Shackleford, D. M.; Morizzi, J.; Mannila, J.; Katneni, K.; Bhamidipati, R.; Zabiulla, K. M.; Joseph, J. T.; Bashyam, S.; Waterson, D.; Witty, M. J.; Hardick, D.; Wittlin, S.; Avery, V.; Charman, S. A.; Chibale, K. 3,5-Diaryl-2aminopyridines as a novel class of orally active antimalarials demonstrating single dose cure in mice and clinical candidate potential. J. Med. Chem. 2012, 55, 3479−3487. (36) Guiguemde, W. A.; Shelat, A. A.; Garcia-Bustos, J. F.; Diagana, T. T.; Gamo, F. J.; Guy, R. K. Global phenotypic screening for antimalarials. Chem. Biol. 2012, 19, 116−129. (37) Plouffe, D.; Brinker, A.; McNamara, C.; Henson, K.; Kato, N.; Kuhen, K.; Nagle, A.; Adrian, F.; Matzen, J. T.; Anderson, P.; Nam, T. G.; Gray, N. S.; Chatterjee, A.; Janes, J.; Yan, S. F.; Trager, R.; Caldwell, J. S.; Schultz, P. G.; Zhou, Y.; Winzeler, E. A. In silico activity profiling reveals the mechanism of action of antimalarials discovered in a highthroughput screen. Proc. Natl. Acad. Sci. U. S. A. 2008, 105, 9059−9064. (38) Holloway, G. A.; Charman, W. N.; Fairlamb, A. H.; Brun, R.; Kaiser, M.; Kostewicz, E.; Novello, P. M.; Parisot, J. P.; Richardson, J.; Street, I. P.; Watson, K. G.; Baell, J. B. Trypanothione reductase highthroughput screening campaign identifies novel classes of inhibitors with antiparasitic activity. Antimicrob. Agents Chemother. 2009, 53, 2824−2833. (39) Berg, S. 127. The search for new trypanocides. Part VII. mAmidinophenyldiazoaminostyrylqyinolinium salts. J. Chem. Soc. 1962, 677−679. (40) Austin, W.; Potter, M.; Taylor, E. Potential trypanocides. The action of polymethylene dihalides on 4-aminoquinaldine. J. Chem. Soc. 1958, 1489−1498. (41) Takasu, K.; Shimogama, T.; Saiin, C.; Kim, H.; Wataya, Y.; Brun, R.; Ihara, M. Synthesis and evaluation of β-carbolinium cations as new antimalarial agents based on π-delocalized lipophilic cation (DLC) hypothesis. Chem. Pharm. Bull. 2005, 53, 653−661. (42) Fulton, J. D.; Joyner, L. P.; King, H.; Osbond, J. M.; Wright, J. Amoebicidal Action and Chemical Constitution. Proc. R. Soc., B 1950, 137, 339−366. (43) List, B. Emil Knoevenagel and the roots of aminocatalysis. Angew. Chem., Int. Ed. Engl. 2010, 49, 1730−1734. (44) Royer, R. 386. The preparation of 4-amino-2-styrylquinoline and some related compounds. J. Chem. Soc. 1949, 1803−1806. 6213

dx.doi.org/10.1021/jm400656s | J. Med. Chem. 2013, 56, 6200−6215

Journal of Medicinal Chemistry

Article

(65) Bulic, B.; Pickhardt, M.; Mandelkow, E.-M.; Mandelkow, E. Tau protein and tau aggregation inhibitors. Neuropharmacology 2010, 59, 276−289. (66) Pudhom, K.; Kasai, K.; Terauchi, H.; Inoue, H.; Kaiser, M.; Brun, R.; Ihara, M.; Takasua, K. Synthesis of three classes of rhodacyanine dyes and evaluation of their in vitro and in vivo antimalarial activity. Bioorg. Med. Chem. 2006, 14, 8550−8563. (67) Hales, D. R.; Welch, A. D. A preliminary study of the anthelmintic activity of cyanine dye #715 in dogs. J. Pharmacol. Exp. Ther. 1953, 107, 310−314. (68) Iglesia, F. A. d. l.; Lake, R. S.; Fitzgerald, J. E.; Bueding, E.; Brusick, D. Bacterial mutagenesis and cell transformation assays of pyrvinium pamoate (povan), an antiparasitic agent. Int. J. Toxicol. 1984, 3, 285− 294. (69) Mehta, R. D.; Hennig, U. G. G.; von Borstel, R. C.; Chatten, L. G. Genetic activity in Saccharomyces cerevisiae and thin-layer chromatographic comparisons of medical grades of pyrvinium pamoate and monopyrvinium salts. Mutat. Res. 1982, 102, 59−69. (70) Esumi, H.; Lu, J.; Kurashima, Y.; Hanaoka, T. Antitumor activity of pyrvinium pamoate, 6-(dimethylamino)-2-[2-(2,5-dimethyl-1-phenyl-1H-pyrrol-3-yl)ethenyl]-1-methyl-quinolinium pamoate salt, showing preferential cytotoxicity during glucose starvation. Cancer Sci. 2004, 95, 685−690. (71) Hilal, H.; Taylor, J. A. Cyanine dyes for the detection of double stranded DNA. J. Biochem. Biophys. Methods 2008, 70, 1104−1108. (72) Egan, T. J.; Hunter, R.; Kaschula, C. H.; Marques, H. M.; Misplon, A.; Walden, J. Structure−function relationships in aminoquinolines: effect of amino and chloro groups on quinoline−hematin complex formation, inhibition of beta-hematin formation, and antiplasmodial activity. J. Med. Chem. 2000, 43, 283−291. (73) Vippagunta, S. R.; Dorn, A.; Matile, H.; Bhattacharjee, A. K.; Karle, J. M.; Ellis, W. Y.; Ridley, R. G.; Vennerstrom, J. L. Structural specificity of chloroquine−hematin binding related to inhibition of hematin polymerization and parasite growth. J. Med. Chem. 1999, 42, 4630−4639. (74) Gorka, A. P.; Alumasa, J. N.; Sherlach, K. S.; Jacobs, L. M.; Nickley, K. B.; Brower, J. P.; de Dios, A. C.; Roepe, P. D. Cytostatic versus cytocidal activities of chloroquine analogues and inhibition of hemozoin crystal growth. Antimicrob. Agents Chemother. 2013, 57, 356− 364. (75) Moura, P. A.; Dame, J. B.; Fidock, D. A. Role of Plasmodium falciparum Digestive Vacuole Plasmepsins in the Specificity and Antimalarial Mode of Action of Cysteine and Aspartic Protease Inhibitors. Antimicrob. Agents Chemother. 2009, 53, 4968−4978. (76) Mungthin, M.; Bray, P. G.; Ridley, R. G.; Ward, S. A. Central role of hemoglobin degradation in mechanisms of action of 4-aminoquinolines, quinoline methanols, and phenanthrene methanols. Antimicrob. Agents Chemother. 1998, 42, 2973−2977. (77) Rosenthal, P. J. Cysteine proteases of malaria parasites. Int. J. Parasitol. 2004, 34, 1489−1499. (78) Hekmat-Nejad, M.; Rathod, P. K. Plasmodium falciparum: kinetic interactions of WR99210 with pyrimethamine-sensitive and pyrimethamine-resistant dihydrofolate reductase. Exp. Parasitol. 1997, 87, 222− 228. (79) Macrae, J. I.; Dixon, M. W.; Dearnley, M. K.; Chua, H. H.; Chambers, J. M.; Kenny, S.; Bottova, I.; Tilley, L.; McConville, M. J. Mitochondrial metabolism of sexual and asexual blood stages of the malaria parasite Plasmodium falciparum. BMC Biol. 2013, 11, 67. (80) Hanssen, E.; Knoechel, C.; Dearnley, M.; Dixon, M. W.; Le Gros, M.; Larabell, C.; Tilley, L. Soft X-ray microscopy analysis of cell volume and hemoglobin content in erythrocytes infected with asexual and sexual stages of Plasmodium falciparum. J. Struct. Biol. 2012, 177, 224−232. (81) Lucantoni, L.; Avery, V. Whole-cell in vitro screening for gametocytocidal compounds. Future Med. Chem. 2012, 4, 2337−2360. (82) Peatey, C. L.; Skinner-Adams, T. S.; Dixon, M. W.; McCarthy, J. S.; Gardiner, D. L.; Trenholme, K. R. Effect of antimalarial drugs on Plasmodium falciparum gametocytes. J. Infect. Dis. 2009, 200, 1518− 1521.

(45) Ruiz, A.; Rocca, P.; Marsais, F.; Godard, A.; Quéguiner, G. Pyridinium chloride: a new reagent for N-demethylation of Nmethylazinium derivatives. Tetrahedron Lett. 1997, 38, 6205−6208. (46) Ho, T.-L. Dealkylation of quaternary ammonium salts with 1,4diazabicyclo[2.2.2]octane. Synthesis 1972, 1972, 702. (47) Wang, M.; Gao, M.; Miller, K. D.; Sledge, G. W.; Hutchins, G. D.; Zheng, Q.-H. Simple synthesis of carbon-11 labeled styryl dyes as new potential PET RNA-specific, living cell imaging probes. Eur. J. Med. Chem. 2009, 44, 2300−2306. (48) Chong, C. R.; Chen, X.; Shi, L.; Liu, J. O.; Sullivan, D. J., Jr. A clinical drug library screen identifies astemizole as an antimalarial agent. Nature Chem. Biol. 2006, 2, 415−416. (49) Bendrat, K.; Berger, B. J.; Cerami, A. Haem polymerization in malaria. Nature 1995, 378, 138−139. (50) Fitch, C. D.; Cai, G. Z.; Chen, Y. F.; Shoemaker, J. D. Involvement of lipids in ferriprotoporphyrin IX polymerization in malaria. Biochim. Biophys. Acta 1999, 1454, 31−37. (51) Jackson, K. E.; Klonis, N.; Ferguson, D. J. P.; Adisa, A.; Dogovski, C.; Tilley, L. Food vacuole-associated lipid bodies and heterogeneous lipid environments in the malaria parasite Plasmodium falciparum. Mol. Microbiol. 2004, 54, 109−122. (52) Sandlin, R. D.; Carter, M. D.; Lee, P. J.; Auschwitz, J. M.; Leed, S. E.; Johnson, J. D.; Wright, D. W. Use of the NP-40 detergent-mediated assay in discovery of inhibitors of beta-hematin crystallization. Antimicrob. Agents Chemother. 2011, 55, 3363−3369. (53) Raynes, K.; Foley, M.; Tilley, L.; Deady, L. W. Novel bisquinoline antimalarials. Synthesis, antimalarial activity, and inhibition of haem polymerisation. Biochem. Pharmacol. 1996, 52, 551−559. (54) Dorn, A.; Vippagunta, S. R.; Matile, H.; Bubendorf, A.; Vennerstrom, J. L.; Ridley, R. G. A comparison and analysis of several ways to promote haematin (haem) polymerisation and an assessment of its initiation in vitro. Biochem. Pharmacol. 1998, 55, 737−747. (55) Krogstad, D. J.; Schlesinger, P. H. The basis of antimalarial action: Non-weak base effects of chloroquine on acid vesicle pH. Am. J. Trop. Med. Hyg. 1987, 36, 213−220. (56) Klonis, N.; Crespo-Ortiz, M. P.; Bottova, I.; Abu-Bakar, N.; Kenny, S.; Rosenthal, P. J.; Tilley, L. Artemisinin activity against Plasmodium falciparum requires hemoglobin uptake and digestion. Proc. Natl. Acad. Sci. U. S. A. 2011, 108, 11405−11410. (57) Crespo, M. P.; Tilley, L.; Klonis, N. Solution behavior of hematin under acidic conditions and implications for its interactions with chloroquine. J. Biol. Inorg. Chem. 2010, 15, 1009−1022. (58) Adjalley, S. H.; Johnston, G. L.; Li, T.; Eastman, R. T.; Ekland, E. H.; Eappen, A. G.; Richman, A.; Sim, B. K.; Lee, M. C.; Hoffman, S. L.; Fidock, D. A. Quantitative assessment of Plasmodium falciparum sexual development reveals potent transmission-blocking activity by methylene blue. Proc. Natl. Acad. Sci. U. S. A. 2011, 108, E1214−E1223. (59) Tomitsuka, E.; Kita, K.; Esumi, H. An anticancer agent, pyrvinium pamoate inhibits the NADH−fumarate reductase systema unique mitochondrial energy metabolism in tumour microenvironments. J. Biochem. 2012, 152, 171−183. (60) Divo, A. A.; Geary, T.; Jensen, J.; Ginsburg, H. The mitochondrion of Plasmodium falciparum visualized by rhodamine 123 fluorescence. J. Protozool. 1985, 32, 442−446. (61) Buchanan, R. A.; Barrow, W. B.; Heffelfinger, J. C.; Kinkel, A. W.; Smith, T. C.; Turner, J. L. Pyrvinium pamoate. Clin. Pharmacol. Ther. 1974, 16, 716−719. (62) Downey, A. S.; Chong, C. R.; Graczyk, T. K.; Sullivan, D. J. Efficacy of pyrvinium pamoate against Cryptosporidium parvum infection in vitro and in a neonatal mouse model. Antimicrob. Agents Chemother. 2008, 52, 3106−3112. (63) Yu, D. H.; Macdonald, J.; Liu, G.; Lee, A. S.; Ly, M.; Davis, T.; Ke, N.; Zhou, D.; Wong-Staal, F.; Li, Q. X. Pyrvinium targets the unfolded protein response to hypoglycemia and its anti-tumor activity is enhanced by combination therapy. PLoS One 2008, 3, e3951. (64) Sheth, U. K. Mechanisms of anthelmintic action. Prog. Drug Res. 1975, 19, 147−157. 6214

dx.doi.org/10.1021/jm400656s | J. Med. Chem. 2013, 56, 6200−6215

Journal of Medicinal Chemistry

Article

(83) Smalley, M. E. Plasmodium falciparum gametocytes: The effect of chloroquine on their development. Trans. R. Soc. Trop. Med. Hyg. 1977, 71, 526−529. (84) Tomitsuka, E.; Kita, K.; Esumi, H. The NADH−fumarate reductase system, a novel mitochondrial energy metabolism, is a new target for anticancer therapy in tumor microenvironments. Ann. N. Y. Acad. Sci. 2010, 1201, 44−49. (85) 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. (86) Foote, S. J.; Thompson, J. K.; Cowman, A. F.; Kemp, D. J. Amplification of the multidrug resistance gene in some chloroquineresistant isolates of P. falciparum. Cell 1989, 57, 921−930. (87) Gardner, M. J.; Hall, N.; Fung, E.; White, O.; Berriman, M.; Hyman, R. W.; Carlton, J. M.; Pain, A.; Nelson, K. E.; Bowman, S.; Paulsen, I. T.; James, K.; Eisen, J. A.; Rutherford, K.; Salzberg, S. L.; Craig, A.; Kyes, S.; Chan, M. S.; Nene, V.; Shallom, S. J.; Suh, B.; Peterson, J.; Angiuoli, S.; Pertea, M.; Allen, J.; Selengut, J.; Haft, D.; Mather, M. W.; Vaidya, A. B.; Martin, D. M.; Fairlamb, A. H.; Fraunholz, M. J.; Roos, D. S.; Ralph, S. A.; McFadden, G. I.; Cummings, L. M.; Subramanian, G. M.; Mungall, C.; Venter, J. C.; Carucci, D. J.; Hoffman, S. L.; Newbold, C.; Davis, R. W.; Fraser, C. M.; Barrell, B. Genome sequence of the human malaria parasite Plasmodium falciparum. Nature 2002, 419, 498−511. (88) Frankland, S.; Adisa, A.; Horrocks, P.; Taraschi, T. F.; Schneider, T.; Elliott, S. R.; Rogerson, S. J.; Knuepfer, E.; Cowman, A. F.; Newbold, C. I.; Tilley, L. Delivery of the malaria virulence protein PfEMP1 to the erythrocyte surface requires cholesterol-rich domains. Eukaryotic Cell 2006, 5, 849−860. (89) Duffy, S.; Avery, V. M. Development and optimization of a novel 384-well anti-malarial imaging assay validated for high-throughput screening. Am. J. Trop. Med. Hyg. 2012, 86, 84−92. (90) Izumiyama, S.; Omura, M.; Takasaki, T.; Ohmae, H.; Asahi, H. Plasmodium falciparum: development and validation of a measure of intraerythrocytic growth using SYBR Green I in a flow cytometer. Exp. Parasitol. 2009, 121, 144−150. (91) Fu, Y.; Tilley, L.; Kenny, S.; Klonis, N. Dual labeling with a far red probe permits analysis of growth and oxidative stress in P. falciparuminfected erythrocytes. Cytometry, Part A 2010, 77, 253−263. (92) Buchanan, M. S.; Davis, R. A.; Duffy, S.; Avery, V. M.; Quinn, R. J. Antimalarial benzylisoquinoline alkaloid from the rainforest tree doryphora sassafras. J. Nat. Prod. 2009, 72, 1541−1543. (93) Kalkanidis, M.; Klonis, N.; Tilley, L.; Deady, L. W. Novel phenothiazine antimalarials: synthesis, antimalarial activity, and inhibition of the formation of β-haematin. Biochem. Pharmacol. 2002, 63, 833−842. (94) Landquist, J. K. 231. Synthetic antimalarials. Part XLVI. Some 4dialkylaminoalkylaminoquinoline derivatives. J. Chem. Soc. 1951, 1038− 1048. (95) Liu, Y.; Ding, Q.; Wu, X. Rearrangement of N,N-di-tertbutoxycarbonylpyridin-4-amines and formation of polyfunctional pyridines. J. Org. Chem. 2008, 73, 6025−6028. (96) Arnold, W. D.; Bounaud, P.; Gosberg, A.; Li, Z.; McDonald, I.; Steensma, R. W.; Wilson, M. E. Pyrrolo-pyridine kinase modulators. U.S. Patent US2006030583 A1, 2006.

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dx.doi.org/10.1021/jm400656s | J. Med. Chem. 2013, 56, 6200−6215