Synthesis, Structure-Activity Relationship, and Antimalarial Efficacy of 6-Chloro-2-arylvinylquinolines

Abstract

There is an urgent need to develop new efficacious antimalarials to address the emerging drug-resistant clinical cases. Our previous phenotypic screening identified styrylquinoline UCF501 as a promising antimalarial compound. To optimize UCF501, we herein report a detailed structure-activity relationship study of 2-arylvinylquinolines, leading to the discovery of potent, low nanomolar antiplasmodial compounds against a Plasmodium falciparum CQ-resistant Dd2 strain, with excellent selectivity profiles (resistance index < 1 and selectivity index > 200). Several metabolically stable 2-arylvinylquinolines are identified as fast-acting agents that kill asexual blood-stage parasites at the trophozoite phase, and the most promising compound 24 also demonstrates transmission blocking potential. Additionally, the monophosphate salt of 24 exhibits excellent in vivo antimalarial efficacy in the murine model without noticeable toxicity. Thus, the 2-arylvinylquinolines represent a promising class of antimalarial drug leads.

Figure 1.

SAR strategy around the quinoline scaffold.

Figure 2.

Stage-specific inhibition of Pf growth by compound 24. Tightly synchronized Dd2 parasites were treated at 6, 18, 30, and 42 HPI with compound 24 at 5 × EC₅₀ concentration. Samples for Giemsa staining and flow cytometry were collected every 12 h following compound addition. (A) Microscopic images of Giemsa-stained thin smears. (B) Histogram plot of YOYO-1-labeled cells from flow cytometric analysis. Results shown are representative of three independent biological replicates.

Figure 3.

Transmission electron micrograph of Pf Dd2 exposed to 5 × EC₅₀ compound 24. (A) Giemsa-stained image of representative parasites processed for TEM. A large vacuole is observed in the cytosol of treated parasites. (B) Micrographs of the parasite after treatment with compound 24 for increasing periods of time. Parasites treated with 0.15% DMSO used as a control. Micrographs of the treated parasite show membrane-bound structures within DV (*), suggesting undigested hemoglobin vesicles. Details of DV from the treated parasite with undigested vesicles (*) and hemozoin crystals are shown in the far-right panel. Scale bar is 0.5 μm.

Figure 4.

Parasitocidal activity of 2-arylvinylquinolines. Asynchronous Dd2 parasite culture was exposed to 5 × EC₅₀ concentrations of test compounds for (A) 6, (B) 12, (C) 24, and (D) 48 h followed by compound removal. Parasite growth was then monitored daily for 96 h. DHA and atovaquone (50 and 6.6 nM) were included as fast- and slow-acting controls, respectively. Parasitemia was determined by microscopy of Giemsa-stained smear. Results shown are representative of three independent biological replicates. Mean ± SEM of three independent readings.

Figure 5.

Activity of compound 24 on 3D7 P. falciparum gametocyte stages. The viability of gametocytes after the exposure of compound 24 was evaluated on early (A) or late (B) gametocyte stages of the 3D7-expressing luciferase parasite. EC₅₀ data represent the means and SEMs of three experiments.

Figure 6.

Effect of the 2-arylvinylaminequinoline derivatives on the β-hematin crystal formation. (A) Images of β-hematin crystals following incubation of 100 μM hemin, propionate buffer, phosphatidylcholine, and varying concentrations of the compound for 16 h at 37 °C. Images were taken using a Nikon Eclipse TE200. (B) Free hemin, indicative of inhibition of β-hematin crystal formation, was determined using a linear calibration curve. Data represent mean ± SEM of three independent experiments.

Figure 7.

Curative property of 2-arylvinylquinoline derivatives. (A) Swiss Webster female mice were infected with P. berghei ANKA luciferase-expressing strain and treated with 25 and 100 mg/kg p.o. once daily 48 h post infection. A total of 7 days after infection, bioluminescence was detected (A) and quantified (B) using an in vivo imaging system (IVIS).

Figure 8.

Effect of compound 24s on the survivability of P. berghei ANKA-infected mice. BALB/c females were infected with a P. berghei ANKA luciferase expressing strain and treated 4 h postinfection with 25 or 100 mg/kg given orally once daily for 4 days. There was no statistically significant difference between the 25 and 100 mg/kg treatment group log-rank (Mantel–Cox) test (p = 0.2636).

Figure 9.

Summary of SAR trends.

Scheme 1.. Synthesis of 6-Substituted 2-Styrylquinolines 8–37^a

^aReagents and conditions: (a) method A for 3a: (i) p-anisidine 1a, ethyl acetoacetate 2, acetic acid, anhydrous magnesium sulfate, and ethanol, 90 °C, 6 h; (ii) Dowtherm, 270 °C, 30 min, 43% for two steps; method B for 3a–d: anilines 1a–d, ethyl acetoacetate 2, PPA, 150 °C, 2 h, 57–68%; (b) phosphorus oxychloride, 105 °C, 2 h, 85–90%; (c) N,N-dimethylaminoalkylamines 5a–b, 130 °C, 24 h, 87–93%; and (d) aromatic aldehydes 7a–l, p-TsNH₂, xylene, 130 °C, 12 h, 60–89%.

Scheme 2.. Synthesis of 6-Substituted 2-Arylvinylquinolines 39–57^a

^aReagents and conditions: (a) p-TsNH₂, aldehydes 38a–k, xylene, 130 °C, 12 h, 48–86%.

Scheme 3.. Synthesis of 6-Substituted 2-Alkylquinolines 58–59^a

^aReagents and conditions: (a) hydrazine hydrate, EtOH, 80 °C, 36 h, 81–83%.

Scheme 4.. Synthesis of 4-Substituted-arylvinylquinolines 62–72^a

^aReagents and conditions: (a) amines 60a–f, 130 °C, EtOH, 12 h, 73–92% and (b) aldehydes 7a, 7h, or 38c, p-TsNH₂, xylene, 130 °C, 12 h, 48–89%.

Scheme 5.. Synthesis of Isonitrile Styrylquinoline 64^a

^aReagents and conditions: (a) triethylphosphite, reflux, 20 h, 88%; (b) Pd/C, H₂, MeOH, overnight, 75%; (c) 50% NaOH, CHCl₃, tetrabutylammonium bromide, DCM, rt to 40 °C, 2 h, 80%; (d) SeO₂, 1,4-dioxane, 80 °C, 6 h, 32%; and (e) potassium tert-butoxide, anhydrous DMF, rt, 1 h, 58%.

Scheme 6.. Synthesis of 4-Aminoarylvinylquinolines 81–87^a

^aReagents and conditions: (a) 2-aminoethanol, EtOH, 130 °C, 48 h, 97%; (b) MsCl, Et₃N, THF, 0 °C, 1 h, 72%; (c) K₂CO₃, amines, anhydrous CH₃CN, reflux, overnight, 80–94%; and (d) aldehydes 7a, 7h, or 38c, p-TsNH₂ xylene, 130 °C, 12 h, 79–91%.

Scheme 7.. Synthesis of 4-Aminoarylvinylquinolines 90–96^a

^aReagents and conditions: (a) aliphatic amines 88a–c, 130 °C, 24 h, 86–93% and (b) aldehydes 7a, 7h, 7j, or 38c, p-TsNH₂, xylene, 130 °C, 12 h, 67–88%.

Scheme 8.. Synthesis of 4-Morpholinobutylaminoarylvinylquinolines 98–114^a

^aReagents and conditions: (a) aldehydes 7i and 97a–p, p-TsNH₂, xylene, 130 °C, 12 h, 52–92%.

Scheme 9.. Synthesis of 4-Morpholinobutylaminoarylquinolines 119–120^a

^aReagents and conditions: (a) ethyl benzoylacetate 115 or ethyl isonicotinoyl acetate 116, 150 °C, PPA, 6 h; (b) phosphorus oxychloride, 105 °C, 2 h, 20–45% for two steps; and (c) 4-morpholinobutanamine, 130 °C, 24 h, 43–49%.