Investigating the Cytsolic Heme Pool in P. falciparum

描述（由申请方提供）：在恶性疟原虫感染疟疾期间，红细胞内寄生虫在专门的食物泡中降解大量血红蛋白。释放的氨基酸被用来支持生长，但血红素主要在食物泡中解毒为疟原虫色素。然而，目前还不清楚有多少血红素分区到寄生虫的胞质溶胶，以及它在寄生虫生理学中的作用。此外，氯喹，一种常用的抗疟疾药物已被建议增加胞质血红素通量，但这尚未得到证实。我们将通过使用函数方法而不是生物系统中容易出现大误差的质量平衡测量来解决这些问题。我们的目标是（1）开发高亲和力的RNA适体血红素使用SELEX，和（2）将它们引入恶性疟原虫，并确定它们对寄生虫生存的影响在氯喹的存在和不存在。由于其负电荷，预期适体是胞质的，并且不应干扰食物泡中的血红素聚合，从而提供了选择性地研究胞质中的血红素池的工具。这些研究对于促进我们了解恶性疟原虫如何处置和/或利用宿主来源的血红素，以及专门研究常用抗疟疾药物发挥其毒性的潜在机制非常重要。对氯喹的耐药性正变得越来越普遍，并通过防止药物在寄生虫内积累的外排机制发生。如果寄生虫的生存与生理性胞质血红素池的状态复杂相关，这可能代表了另一种药物开发靶点，可以绕过已知的耐药机制。

DESCRIPTION (provided by applicant): During malarial infection by Plasmodium falciparum, significant, amounts of hemoglobin are degraded by the intraerythrocytic parasite in a specialized food vacuole. The released amino acids are utilized to support growth, but heme is mostly detoxified to hemozoin in the food vacuole. However, it is unclear how much heme partitions to the parasite's cytosol, and the role it plays in parasite physiology. Furthermore, chloroquine, a commonly used anti-malarial drug has been suggested to increase cytosolic heme flux, but this has not been proven. We will address these issues by using a functional approach rather than mass balance measurements that are prone to large errors in biological systems. Our goals are to (1) develop high affinity RNA aptamers to heme using SELEX, and, (2) introduce them into P. falciparum, and determine their impact on parasite survival in the presence and absence of chloroquine. The aptamers are expected to be cytosolic given their negative charge, and should not interfere with heme polymerization in the food vacuole, thus providing a tool to selectively investigate the heme pool in the cytosol. These studies are important to advance our understanding of how P. falciparum disposes and/or utilizes host-derived heme, and to specifically investigate a potential mechanism by which the commonly used anti-malarial drugs exert their toxicity. Resistance to chloroquine is becoming increasingly widespread, and occurs via efflux mechanisms that prevent drug accumulation within the parasite. If parasite survival is intricately associated with the status of a physiologic cytosolic heme pool, this may represent another drug development target that circumvents the known resistance mechanism.