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Inhibition of Hemozoin Formation in Plasmodium falciparum Trophozoite Extracts by Heme Analogs: Possible Implication in the Resistance to Malaria Conferred by the β-Thalassemia Trait

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Abstract

Background

Human falciparum malaria, caused by the intracellular protozoa Plasmodium falciparum, results in 1–2 million deaths per year. P. falciparum digests host erythrocyte hemoglobin within its food vacuole, resulting in the release of potentially toxic free heme. A parasite-specific heme polymerization activity detoxifies the free heme by cross-linking the heme monomers to form hemozoin or malaria pigment. This biochemical process is the target of the widely successful antimalarial drug chloroquine, which is rapidly losing its effectiveness due to the spread of chloroquine resistance. We have shown that chloroquine resistance is not due to changes in the overall catalytic activity of heme polymerization or its chloroquine sensitivity. Therefore, the heme polymerization activity remains a potential target for novel antimalarials. In this study, we investigated the ability of heme analogs to inhibit heme polymerization and parasite growth in erythrocytes.

Materials and Methods

Incorporation of radioactive hemin substrate into an insoluble hemozoin pellet was used to determine heme polymerization. Incorporation of radioactive hypoxanthine into the nucleic add of dividing parasites was used to determine the effects of heme analogs on parasite growth. Microscopic and biochemical measurements were made to determine the extent of heme analog entry into infected erythrocytes.

Results

The heme analogs tin protoporphyrin IX (SnPP), zinc protoporphyrin IX (ZnPP), and zinc deuteroporphyrin IX, 2,4 bisglycol (ZnBG) inhibited polymerization at micromolar concentrations (ZnPP ≪ SnPP < ZnBG). However, they did not inhibit parasite growth since they failed to gain access to the site of polymerization, the parasite’s food vacuole. Finally, we observed high ZnPP levels in erythrocytes from two patients with β-thalassemia trait, which may inhibit heme polymerization.

Conclusions

The heme analogs tested were able to inhibit hemozoin formation in Plasmodium falciparum trophozite extracts. The increased ZnPP levels found in thalassemic erythrocytes suggest that these may contribute, at least in part, to the observed antimalarial protection conferred by the β-thalassemia trait. This finding may lead to the development of new forms of antimalarial therapy.

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Acknowledgments

We want to thank Gary Latortue for technical help and Dr. Patricia Griadino at the Pediatric Hematology Department of New York Hospital for the β-thalassemia blood samples. This work was supported by National Institutes of Health Grant RO1-AI-30660-04, Minority Individual in Postdoctoral Training Supplement Grant RO1-AI-30660-SI (JAM) and the UNDP/World Bank/WHO Special Program for Research in Tropical Diseases (AFGS).

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Authors and Affiliations

  1. The Picower Institute for Medical Research, 350 Community Drive, Manhasset, New York, 11030, USA

    James A. Martiney, Anthony Cerami & Andrew F. G. Slater

  2. Division of Toxicology, The Karolinksa Institute, Stockholm, Sweden

    Andrew F. G. Slater

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  1. James A. Martiney
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  2. Anthony Cerami
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  3. Andrew F. G. Slater
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Martiney, J.A., Cerami, A. & Slater, A.F.G. Inhibition of Hemozoin Formation in Plasmodium falciparum Trophozoite Extracts by Heme Analogs: Possible Implication in the Resistance to Malaria Conferred by the β-Thalassemia Trait. Mol Med 2, 236–246 (1996). https://doi.org/10.1007/BF03401620

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