Atovaquone-Proguanil in Combination With Artesunate to Treat Multidrug-Resistant P. falciparum Malaria in Cambodia: An Open-Label Randomized Trial

Abstract

Background Recent artemisinin-combination therapy failures in Cambodia prompted a search for alternatives. Atovaquone‐proguanil (AP), a safe, effective treatment for multidrug-resistant Plasmodium falciparum (P.f.), previously demonstrated additive effects in combination with artesunate (AS). Methods Patients with P.f. or mixed-species infection (n = 205) in Anlong Veng (AV; n = 157) and Kratie (KT; n = 48), Cambodia, were randomized open-label 1:1 to a fixed‐dose 3-day AP regimen +/-3 days of co‐administered artesunate (ASAP). Single low-dose primaquine (PQ, 15 mg) was given on day 1 to prevent gametocyte-mediated transmission. Results Polymerase chain reaction–adjusted adequate clinical and parasitological response at 42 days was 90% for AP (95% confidence interval [CI], 82%–95%) and 92% for ASAP (95% CI, 83%–96%; P = .73). The median parasite clearance time was 72 hours for ASAP in AV vs 56 hours in KT (P < .001) and was no different than AP alone. At 1 week postprimaquine, 7% of the ASAP group carried microscopic gametocytes vs 29% for AP alone (P = .0001). Nearly all P.f. isolates had C580Y K13 propeller artemisinin resistance mutations (AV 99%; KT 88%). Only 1 of 14 treatment failures carried the cytochrome bc1 (Pfcytb) atovaquone resistance mutation, which was not present at baseline. P.f. isolates remained atovaquone sensitive in vitro but cycloguanil resistant, with a triple P.f. dihydrofolate reductase mutation. Conclusions Atovaquone-proguanil remained marginally effective in Cambodia (≥90%) with minimal Pfcytb mutations observed. Treatment failures in the presence of ex vivo atovaquone sensitivity and adequate plasma levels may be attributable to cycloguanil and/or artemisinin resistance. Artesunate co-administration provided little additional blood-stage efficacy but reduced post-treatment gametocyte carriage in combination with AP beyond single low-dose primaquine.

Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

“…Multiple copies of PM2 gene might help overcome the fitness cost of PfCRT variants by increasing the rate of hemoglobin degradation, leading to quick sequestration of reactive heme into hemozoin (32).…”

“…All P. falciparum samples in this study were collected in Cambodia under the therapeutic efficacy of atovaquone (ATQ)-proguanil (PG) and artesunate (AS)-ATQ-PG for uncomplicated P. falciparum malaria, in areas of documented multidrug resistance (Clinicaltrials.gov NCT02297477) [32]. The P. falciparum samples were collected on Day 0 (screening) and then cryopreserved.…”

“…Initially, 26 parasites were selected from the 88 clinical isolates based on the availability of ex vivo drug sensitivity, ex vivo molecular markers and parasitaemia of ≥ 0.5%. [32]. After two attempts of thawing process, seventeen (65.4%) P. falciparum clinical isolates were successfully culture-adapted.…”

“…Of 221 isolates available for molecular marker testing, all samples held a single copy number of pfmdr1, 211 samples (95.5%) contained C580Y K13 propeller artemisinin resistance mutations, and 184 samples (83.3%) and 37 samples (16.7%) harbored multiple copies and single copy of PM2, respectively [32]. Table 1 illustrates the molecular genotypes of 17 culture-adapted isolates.…”

“…To better understand the main factors leading to PPQ resistant phenotypes, 17 parasite isolates collected as part of a therapeutic efficacy study (Clinicaltrials.gov NCT02297477) in Cambodia [32] were successfully culture-adapted. They were analyzed for copy number variation of pfmdr1, PM2 and SNPs of K13, Exo, and PfCRT, and correlated with survival assays and drug susceptibility phenotypes.…”