Antimalarial efficacy in Plasmodium berghei-Infected Mice

Mice represent an ideal rodent model for Plasmodium berghei infection, which is fatal for these animals [39]. This experimental model simulates the human infection produced by Plasmodium falciparum, which is also lethal in non-treated humans. We used two treatment protocols to determine ATM efficacy in the mouse model. The first one was applied early after mice infection with high doses (total 40 mg and 80 mg/kg divided into 4 daily doses) following the “Four-day test” protocol [13,39]. This protocol aims to evidence efficacy and any general toxicity of the formulations in infected animals in repeated dose experiments. Efficacy was evaluated against chloroquine-sensitive P. berghei NK65-infected mice.

The second efficacy protocol consisted of the treatment of infected mice with established infection (15% blood parasitemia) with a single low dose of ATM (20 mg/kg IV) to better distinguish the formulation profile of the efficacy following time. An infective inoculum was prepared from an infected donor mouse with rising parasitemia (20%). Swiss female mice (18–22 g) were infected (IV) on day zero with 1 × 10⁶ P. berghei parasitized red blood cells (RBC) in 0.2 mL of phosphate-buffered saline and randomly divided into groups. Then, the animals were treated on day 2, with only one IV dose of 20 mg/kg free-ATM solution or ATM-PCL-NCs. Control groups received only ATM vehicles or blank PCL NCs (blank-NCs). Thin blood smears were prepared with blood collected from the tail vein of all animals on days 3, 5, 7, 9, 14, 25, and 60 after infection. Parasitemia was measured in Giemsa-stained smears, counting at least 3000 RBC to determine the percentage of infected ones.