INHIBITION OF HEMOZOIN FORMATION IN PLASMODIUM-FALCIPARUM TROPHOZOITEEXTRACTS BY HEME ANALOGS - POSSIBLE IMPLICATION IN THE RESISTANCE TO MALARIA CONFERRED BY THE BETA-THALASSEMIA TRAIT

Background: Human falciparum malaria, caused by the intracellular prot ozoa Plasmodium falciparum, results in 1-2 million deaths per year. P. falciparum digests host erythrocyte hemoglobin within its food vacuol e, resulting in the release of potentially toxic free heme. A parasite -specific heme polymerization activity detoxifies the free heme by cro ss-linking the heme monomers to form hemozoin or malaria pigment. This biochemical process is the target of the widely successful antimalari al drug chloroquine, which is rapidly losing its effectiveness due to the spread of chloroquine resistance. We have shown that chloroquine r esistance is not due to changes in the overall catalytic activity of h eme polymerization or its chloroquine sensitivity. Therefore, the heme polymerization activity remains a potential target for novel antimala rials. In this study, we investigated the ability of heme analogs to i nhibit heme polymerization and parasite growth in erythrocytes. Materi als and Methods: Incorporation of radioactive hemin substrate into an insoluble hemozoin pellet was used to determine heme polymerization. I ncorporation of radioactive hypoxanthine into the nucleic acid of divi ding parasites was used to determine the effects of heme analogs on pa rasite growth. Microscopic and biochemical measurements were made to d etermine the extent of heme analog entry into infected erythrocytes. R esults: The heme analogs tin protoporphyrin IX (SnPP), zinc protoporph yrin IX (ZnPP), and zinc deuteroporphyrin IX, 2,4 bisglycol (ZnBG) inh ibited polymerization at micromolar concentrations (ZnPP much less tha n SnPP < ZnBG). However, they did not inhibit parasite growth since th ey failed to gain access to the site of polymerization, the parasite's food vacuole. Finally, we observed high ZnPP levels in erythrocytes f rom two patients with beta-thalassemia trait, which may inhibit heme p olymerization. Conclusions: The heme analogs tested were able to inhib it hemozoin formation in Plasmodium falciparum trophozite extracts. Th e increased ZnPP levels found in thalassemic erythrocytes suggest that these may contribute al least in part, to the observed antimalarial p rotection conferred by the beta-thalassemia trait. This finding may le ad to the development of new forms of antimalarial therapy.