Atovaquone-Proguanil Remains a Potential Stopgap Therapy for Multidrug-Resistant Plasmodium falciparum in Areas along the Thai-Cambodian Border

Abstract

Our recent report of dihydroartemisinin-piperaquine failure to treat Plasmodium falciparum infections in Cambodia adds new urgency to the search for alternative treatments. Despite dihydroartemisinin-piperaquine failure, and higher piperaquine 50% inhibitory concentrations (IC50s) following reanalysis than those previously reported, P. falciparum remained sensitive to atovaquone (ATQ) in vitro. There were no point mutations in the P. falciparum cytochrome b ATQ resistance gene. Mefloquine, artemisinin, chloroquine, and quinine IC50s remained comparable to those from other recent reports. Atovaquone-proguanil may be a useful stopgap but remains susceptible to developing resistance when used as blood-stage therapy.

FIG 1

Ex vivo drug susceptibility of P. falciparum isolates from Cambodia. IC₅₀s (in nanomoles) of P. falciparum monoinfection are plotted for each drug, with their geometric mean indicated by a red bar, and indicated by the value below each cluster of data points. The dashed lines denote geometric mean IC₅₀s against the P. falciparum W2 (green) and C2B (blue) reference clones. Red circles indicate isolates that were not originally cleared by 674 nM PPQ.

FIG 2

PPQ dose-response curves for 3 illustrative P. falciparum isolates that were not cleared at the maximum PPQ concentration used in the original assay. The colors denote different isolates. The solid lines are growth inhibition curves interpolated by HRP-2 OD, as measured by serial PPQ dilution with concentrations ranging from 0 to 674 nM. The dashed lines represent reanalyzed curves by including the control OD value for 100% HRP-2 inhibition (2,000 ng/ml CQ) and refitting the curve. This was equivalent to a concentration of 53,905 nM PPQ (denoted by closed circles); reinterpolated IC₅₀s using these values are denoted by vertical colored lines for each isolate.

FIG 3

DNA sequencing analysis for cytb mutations revealed all of the 108 isolates to be wild-type, despite 2 P. falciparum isolates positive for mutations in the HRM-PCR. Y268C, N and S are previously described SNP associated with atovaquone resistance. While the 3D7 P. falciparum clone shares the wild-type sequence, the C2B and TM90 clones share the Y268N SNP.