High-Content Screening of MMV Pathogen Box for Plasmodium falciparum Digestive Vacuole Disrupting Molecules Reveals Valuable Starting Points for Drug Discovery [PublishAheadOfPrint]

Plasmodium falciparum infections leading to malaria have severe clinical manifestations and high mortality rates. Chloroquine (CQ), a former mainstay of malaria chemotherapy, has been rendered ineffective due to the emergence of wide-spread resistance. Recent studies, however, have unveiled a novel mode of action in which low micromolar levels of CQ permeabilized the parasite's digestive vacuole (DV) membrane, leading to calcium efflux, mitochondrial depolarization and DNA degradation. These phenotypes implicate the DV as an alternative target of CQ and suggests that DV disruption is an attractive target for exploitation through the screening for DV-disruptive antimalarials. In the current study, high-content screening was performed on the Medicines for Malaria Venture (MMV) 'Pathogen Box' (2015) to select for compounds which disrupted the DV membrane as measured by the leakage of intravacuolar Ca²⁺ using the calcium probe Fluo-4 AM. The hits were further characterized by hemozoin biocrystallization inhibition assays and dose-response IC₅₀ assays across resistant and sensitive strains. Three hits - MMV676380, MMV085071 and MMV687812 were shown to demonstrate lack of CQ cross-resistance in parasite strains and field isolates. Through systematic analyses, MMV085071 emerged as the top hit due to its rapid parasiticidal effect, low nanomolar IC₅₀ and good efficacy in triggering DV disruption, mitochondrial degradation and DNA fragmentation in P. falciparum. These programmed cell death (PCD)-like phenotypes following permeabilization of the DV suggests that these compounds kill the parasite by a PCD-like mechanism. From the drug development perspective, the identification of MMV085071 as a potent DV disruptor offers a promising starting point for subsequent hit-to-lead generation and optimization through structure-activity relationships.

http://ift.tt/2AI4Yb4