IRON METABOLISM IN PLASMODIUM

抗疟疾耐药性的增加是目前恶性疟疾死亡率不断上升的一个重要因素。目前使用的两大类抗疟疾药物，如氯喹等喹啉类药物和青蒿素等内源性过氧化物质，会干扰铁的新陈代谢，从而杀死寄生虫。铁络合剂虽然对细菌和哺乳动物细胞具有细胞抑制作用，但实际上可以杀死疟原虫，尽管寄生的红细胞中含有毫米级的血红素铁。这表明生物可利用的铁的数量是有限的，对寄生虫的生存至关重要。因此，干扰铁代谢是疟原虫重要且有前景的化疗靶点。这项提议的长期目标是通过进一步定义寄生虫铁的重要来源和表征铁向基本间隔的运输来改进有针对性的抗疟疾化疗。初步数据显示，红细胞铁蛋白是铁的来源之一。最近克隆的恶性疟原虫与哺乳动物铁转运蛋白天然抗性相关巨噬细胞蛋白(NRAMP)的同源物将能够表征寄生虫的铁运输。其具体目的是测试红细胞铁作为铁源，定位PfNRAMP所在的隔室，评估PfNRAMP运输二价金属阳离子的特异性，评估这种质子/阳离子转运体的动力学和抑制作用，并分析其在喹啉敏感和耐药寄生虫中的表达水平。为了达到这些目的，我们将采用在封闭的红细胞中培养疟原虫、重组基因表达、免疫定位、细胞转染和定点诱变等技术。这项研究的重大影响是提高了对抗疟疾药物干扰的疟原虫铁代谢的理解。为美国军方和旅行者以及流行人群改善目前的抗疟疾设施是疟疾控制的一个重要组成部分。

An increase in antimalarial drug resistance is an important contributing component to the current escalating mortality due to Plasmodium falciparum malaria. Two major classes of currently used antimalarials, the quinolines like chloroquine and the endoperoxides like artmesinin, interfere with iron metabolism to kill the parasite. Iron chelators, while cytostatic for bacterial and mammalian cells, actually kill Plasmodium despite the availability of millimolar heme iron in the parasitized erythrocyte. This indicates that the amount of bioavailable iron is limited and crucial for parasite survival. Therefore, interference with iron metabolism is an important and promising chemotherapeutic target for Plasmodium. The long term objectives of this proposal are to improve targeted antimalarial chemotherapy by further definition of the vital sources of parasitic iron and by characterization of iron transport to essential compartments. Preliminary data implicate erythrocytic ferritin as a source of iron. Recent cloning of the P. falciparum homologue to the mammalian iron transporter Natural Resistance Associated Macrophage Protein (NRAMP) will enable characterization of parasite iron transport. The specific aims are to test erythrocytic iron as an iron source, to localize the compartments in which PfNRAMP resides, to assess the specificity of divalent metal cations transported by PfNRAMP, to evaluate the kinetics and inhibition of this proton/cation symporter and to analyze its level of expression amongst quinoline-sensitive and -resistant parasites. The techniques of Plasmodium culture in resealed erythrocytes, recombinant gene expression, immunolocalization, cell transfection, and site-directed mutagenesis will be used to achieve these aims. The significant impact of this research is to improve the understanding of Plasmodium iron metabolism that is perturbed by antimalarial drugs. To improve the present antimalarial armamentarium for the US military and travelers as well as endemic populations is an essential component of malarial control.