High Target Homology Does Not Guarantee Inhibition: Aminothiazoles Emerge as Inhibitors of Plasmodium falciparum.

In this study, we identified three novel compound classes with potent activity against Plasmodium falciparum, the most dangerous human malarial parasite. Resistance of this pathogen to known drugs is increasing, and compounds with different modes of action are urgently needed. One promising drug target is the enzyme 1-deoxy-d-xylulose-5-phosphate synthase (DXPS) of the methylerythritol 4-phosphate (MEP) pathway for which we have previously identified three active compound classes against Mycobacterium tuberculosis. The close structural similarities of the active sites of the DXPS enzymes of P. falciparum and M. tuberculosis prompted investigation of their antiparasitic action, all classes display good cell-based activity. Through structure-activity relationship studies, we increased their antimalarial potency and two classes also show good metabolic stability and low toxicity against human liver cells. The most active compound 1 inhibits the growth of blood-stage P. falciparum with an IC50 of 600 nM. The results from three different methods for target validation of compound 1 suggest no engagement of DXPS. All inhibitor classes are active against chloroquine-resistant strains, confirming a new mode of action that has to be further investigated.

Targeting the Plasmodium falciparum plasmepsin V by ligand-based virtual screening.

Malaria is a devastating disease depending only on chemotherapy as treatment. However, medication is losing efficacy, and therefore, there is an urgent need for the discovery of novel pharmaceutics. Recently, plasmepsin V, an aspartic protease anchored in the endoplasmaic reticulum, was demonstrated as responsible for the trafficking of parasite-derived proteins to the erythrocytic surface and further validated as a drug target. In this sense, ligand-based virtual screening has been applied to design inhibitors that target plasmepsin V of P. falciparum (PMV). After screening 5.5 million compounds, four novel plasmepsin inhibitors have been identified which were subsequently analyzed for the potency at the cellular level. Since PMV is membrane-anchored, the verification in vivo by using transgenic PMV overexpressing P. falciparum cells has been performed in order to evaluate drug efficacy. Two lead compounds, revealing IC50 values were 44.2 and 19.1 µm, have been identified targeting plasmepsin V in vivo and do not significantly affect the cell viability of human cells up to 300 µm. We herein report the use of the consensus of individual virtual screening as a new technique to design new ligands, and we propose two new lead compounds as novel protease inhibitors to target malaria.

Plasmodium falciparum GFP-E-NTPDase expression at the intraerythrocytic stages and its inhibition blocks the development of the human malaria parasite.

Plasmodium falciparum is the causative agent of the most dangerous form of malaria in humans. It has been reported that the P. falciparum genome encodes for a single ecto-nucleoside triphosphate diphosphohydrolase (E-NTPDase), an enzyme that hydrolyzes extracellular tri- and di-phosphate nucleotides. The E-NTPDases are known for participating in invasion and as a virulence factor in many pathogenic protozoa. Despite its presence in the parasite genome, currently, no information exists about the activity of this predicted protein. Here, we show for the first time that P. falciparum E-NTPDase is relevant for parasite lifecycle as inhibition of this enzyme impairs the development of P. falciparum within red blood cells (RBCs). ATPase activity could be detected in rings, trophozoites, and schizonts, as well as qRT-PCR, confirming that E-NTPDase is expressed throughout the intraerythrocytic cycle. In addition, transfection of a construct which expresses approximately the first 500 bp of an E-NTPDase-GFP chimera shows that E-NTPDase co-localizes with the endoplasmic reticulum (ER) in the early stages and with the digestive vacuole (DV) in the late stages of P. falciparum intraerythrocytic cycle.