Additive In Vitro Antiplasmodial Effect of N-Alkyl and N-Benzyl-1,10-Phenanthroline Derivatives and Cysteine Protease Inhibitor E64

Potential new targets for antimalarial chemotherapy include parasite proteases, which are required for several cellular functions during the Plasmodium falciparum life cycle. Four new derivatives of N-alkyl and N-benzyl-1,10-phenanthroline have been synthesized. Those are (1)-N-methyl-1,10-phenanthrolinium sulfate, (1)-N-ethyl-1,10-phenanthrolinium sulfate, (1)-N-benzyl-1,10-phenanthrolinium chloride, and (1)-N-benzyl-1,10-phenanthrolinium iodide. Those compounds had potential antiplasmodial activity with IC50 values from 260.42 to 465.38 nM. Cysteine proteinase inhibitor E64 was used to investigate the mechanism of action of N-alkyl and N-benzyl-1,10-phenanthroline derivatives. A modified fixed-ratio isobologram method was used to study the in vitro interactions between the new compounds with either E64 or chloroquine. The interaction between N-alkyl and N-benzyl-1,10-phenanthroline derivatives and E64 was additive as well as their interactions with chloroquine were also additive. Antimalarial mechanism of chloroquine is mainly on the inhibition of hemozoin formation. As the interaction of chloroquine and E64 was additive, the results indicated that these new compounds had a mechanism of action by inhibiting Plasmodium proteases. 1. Introduction The erythrocytic life cycle of Plasmodium, which is responsible for all clinical manifestations of malaria, begins when free merozoites invade erythrocytes. The intraerythrocytic parasites develop from small ring-stage organisms to larger, more metabolically active trophozoites and then to multinucleated schizonts. The erythrocytic cycle is completed when mature schizonts rupture erythrocytes, releasing numerous invasive merozoites. Proteases appear to be required for cleavage of red blood cell ankyrin to facilitate host cell rupture and subsequent reinvasion of erythrocytes by merozoites, and for the degradation of hemoglobin ingested by intraerythrocytic trophozoites [1, 2]. Extensive evidence suggest that the degradation of hemoglobin is necessary for the growth of erythrocytic malaria parasite, apparently to provide free amino acids for parasite protein synthesis [3, 4]. In P. falciparum, hemoglobin degradation occurs predominantly in trophozoites and early schizonts, the stages at which the parasites are most metabolically active. Trophozoite ingest erythrocyte cytoplasm and transport it to a large central food vacuole. In the food vacuole, intact heme is released from hemoglobin to form the major component of malarial hemozoin pigment [5]. The food vacuole appears to be the site of action of a number of