Real-Time Imaging of Antimalarial Drug Effects on Plasmodium falciparum-Infected Red Blood Cells

Malaria remains a significant global health challenge, exacerbated by emerging drug resistance, which has been recurring ever since Chloroquine resistance was reported to spread in the 1960s. Cabamiquine (M5717), a promising antimalarial candidate, targets Plasmodium falciparum elongation factor 2 (PfeEF2) and has demonstrated high potency and multi-stage activity. However, differences in parasite-killing kinetics require further exploration. This proof-of-concept study employs time-lapse microscopy to assess the effects of Chloroquine and Cabamiquine on P. falciparum-infected red blood cells. Using a fluorogenic DNA dye, we monitored morphological changes and cell death at the single-cell level. Unlike the faster-killing Chloroquine, our findings confirm a lag phase of the effect of Cabamiquine and reveal a delay before host cell lysis. This delayed cytotoxic effect underscores the cytostatic phase of the drug prior to killing and highlights its potential in combination therapies. Our novel imaging approach provides a comprehensive fingerprinting method to quantify cytostatic, cytotoxic, and cytolytic activities of antimalarial drugs, offering valuable insights into their mechanisms of action at a cellular level therefore enabling to discriminate the steps prior to parasite clearance in hosts.