Overexpression of plasmepsin II and plasmepsin III does not directly cause reduction in Plasmodium falciparum sensitivity to artesunate, chloroquine and piperaquine

Duangkamon Loesbanluechai,Namfon Kotanan,Cristina De Cozar,Theerarat Kochakarn,Megan R Ansbro,Kesinee Chotivanich,Nicholas J White,Prapon Wilairat,Marcus C.S Lee,Francisco Javier Gamo,Laura Maria Sanz,Thanat Chookajorn,Krittikorn Kümpornsin

doi:10.1016/j.ijpddr.2018.11.004

Abstract

Artemisinin derivatives and their partner drugs in artemisinin combination therapies (ACTs) have played a pivotal role in global malaria mortality reduction during the last two decades. The loss of artemisinin efficacy due to evolving drug-resistant parasites could become a serious global health threat. Dihydroartemisinin-piperaquine is a well tolerated and generally highly effective ACT. The implementation of a partner drug in ACTs is critical in the control of emerging artemisinin resistance. Even though artemisinin is highly effective in parasite clearance, it is labile in the human body. A partner drug is necessary for killing the remaining parasites when the pulses of artemisinin have ceased. A population of Plasmodium falciparum parasites in Cambodia and adjacent countries has become resistant to piperaquine. Increased copy number of the genes encoding the haemoglobinases Plasmepsin II and Plasmepsin III has been linked with piperaquine resistance by genome-wide association studies and in clinical trials, leading to the use of increased plasmepsin II/plasmepsin III copy number as a molecular marker for piperaquine resistance. Here we demonstrate that overexpression of plasmepsin II and plasmepsin III in the 3D7 genetic background failed to change the susceptibility of P. falciparum to artemisinin, chloroquine and piperaquine by both a standard dose-response analysis and a piperaquine survival assay. Whilst plasmepsin copy number polymorphism is currently implemented as a molecular surveillance resistance marker, further studies to discover the molecular basis of piperaquine resistance and potential epistatic interactions are needed.

Highlights

Intraerythrocytic malaria parasites propagate successfully inside the human host by devouring a vast amount of haemoglobin
P. falciparum 3D7 parasites were transfected with malarial expression vectors containing plasmepsin II and plasmepsin III with the empty vector as a negative control
We present the evidence that the overexpression of plasmepsin II and plasmepsin III alone does not affect P. falciparum susceptibility to artesunate, chloroquine or piperaquine

Summary

Introduction

Intraerythrocytic malaria parasites propagate successfully inside the human host by devouring a vast amount of haemoglobin. This is achieved by digesting the haemoglobin proteins by proteases in the food vacuole (Francis et al, 1997b). The evolutionary history of these gene families reveals their specialized roles in the Plasmodium species as reflected by their respective independent gene expansion events (Ponsuwanna et al, 2016). This protein complex is associated with the actions of several antimalarial drugs including the quinoline related antimalarials (quinine, chloroquine, mefloquine etc) and artemisinin (Chugh et al, 2013). Genetic variations in these genes are associated with changes in drug susceptibility (Amato et al, 2017; Klonis et al, 2011; Witkowski et al, 2017)

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