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Assessing drug efficacy against Plasmodium falciparum liver stages in vivo
Erika L. Flannery, … , Stefan H.I. Kappe, Sebastian A. Mikolajczak
Erika L. Flannery, … , Stefan H.I. Kappe, Sebastian A. Mikolajczak
Published January 11, 2018
Citation Information: JCI Insight. 2018;3(1):e92587. https://doi.org/10.1172/jci.insight.92587.
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Resource and Technical Advance Infectious disease Microbiology

Assessing drug efficacy against Plasmodium falciparum liver stages in vivo

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Abstract

Malaria eradication necessitates new tools to fight the evolving and complex Plasmodium pathogens. These tools include prophylactic drugs that eliminate Plasmodium liver stages and consequently prevent clinical disease, decrease transmission, and reduce the propensity for resistance development. Currently, the identification of these drugs relies on in vitro P. falciparum liver stage assays or in vivo causal prophylaxis assays using rodent malaria parasites; there is no method to directly test in vivo liver stage activity of candidate antimalarials against the human malaria–causing parasite P. falciparum. Here, we use a liver-chimeric humanized mouse (FRG huHep) to demonstrate in vivo P. falciparum liver stage development and describe the efficacy of clinically used and candidate antimalarials with prophylactic activity. We show that daily administration of atovaquone-proguanil (ATQ-PG; ATQ, 30 mg/kg, and PG, 10 mg/kg) protects 5 of 5 mice from liver stage infection, consistent with the use in humans as a causal prophylactic drug. Single-dose primaquine (60 mg/kg) has similar activity to that observed in humans, demonstrating the activity of this drug (and its active metabolites) in FRG huHep mice. We also show that DSM265, a selective Plasmodial dihydroorotate dehydrogenase inhibitor with causal prophylactic activity in humans, reduces liver stage burden in FRG huHep mice. Finally, we measured liver stage–to–blood stage transition of the parasite, the ultimate readout of prophylactic activity and measurement of infective capacity of parasites in the liver, to show that ATQ-PG reduces blood stage patency to below the limit of quantitation by quantitative PCR (qPCR). The FRG huHep model, thus, provides a platform for preclinical evaluation of drug candidates for liver stage causal prophylactic activity, pharmacokinetic/pharmacodynamics studies, and biological studies to investigate the mechanism of action of liver stage active antimalarials.

Authors

Erika L. Flannery, Lander Foquet, Vorada Chuenchob, Matthew Fishbaugher, Zachary Billman, Mary Jane Navarro, William Betz, Tayla M. Olsen, Joshua Lee, Nelly Camargo, Thao Nguyen, Carola Schafer, Brandon K. Sack, Elizabeth M. Wilson, Jessica Saunders, John Bial, Brice Campo, Susan A. Charman, Sean C. Murphy, Margaret A. Phillips, Stefan H.I. Kappe, Sebastian A. Mikolajczak

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Figure 1

P.falciparum liver infection in FRG huHep mice is established by mosquito bite and prevented with atovaquone-proguanil.

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P.falciparum liver infection in FRG huHep mice is established by mosqui...
(A) Human GAPDH transcript levels measured by qPCR in duplicate in 7 different FRG huHep mice. M, mouse; NTC, no template control. (B) Human albumin levels measured prior to FRG huHep infection. (C) P. falciparum NF54HT-GFP-luc liver stage burden 6 days after infection measured by in vivo immunoflourescence (IVIS) or 18S rRNA qPCR. Mice were infected with either 1 × 106 freshly dissected sporozoites by i.v. injection or the direct feeding of 50 infected mosquitoes. Error bars for qPCR measurements represent mean ± SEM. (D) Correlation of liver burden estimates using qPCR and IVIS. Pearson correlation r2 = 0.53, P < 0.001. Dashed lines denote upper and lower 95% CI. (E) Radiance levels 6 days after infection after treatment with atovaquone-proguanil (ATQ-PG) from day –1 to day 6. Mice were infected by the bite of 50 infectious mosquitoes. Unpaired 2-tailed Student’s t test, ****P < 0.0001. (F) IVIS images of luminescence in livers of live FRG huHep mice from (E). Each symbol represents 1 mouse (B–E). Dashed line represents background radiance (C and E).

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