Discovery of chemically validated malaria liver stage targets

描述（由申请人提供）：由于血液期感染产生疟疾的大多数临床表现，因此药物开发主要集中于开发针对恶性疟原虫（最严重形式的人类疟疾的病原体）的杀疟原虫剂。资金的增加和对寄生虫抗药性问题的日益认识有助于推动一些新的杀寄生虫剂进入开发管道，甚至进入临床试验。另一方面，这些候选药物中很少有对疟疾红细胞外期有效的，甚至更少的药物可能提供对间日疟原虫疟疾的根治。随着消灭疟疾成为世界卫生界的一个更高优先事项，对可替代伯氨喹的药物的需求甚至更为迫切。为了刺激针对肝脏阶段的药物开发活性，需要更多的工作来了解肝脏阶段生物学，并发现其活性对肝脏和血液阶段开发都至关重要的靶标。我将使用化学遗传学方法来研究对血液和肝脏寄生虫发育至关重要的途径，然后描述这种方法揭示的靶标。具体来说，我将在亚致死浓度的小分子下培养寄生虫，这些小分子对血液和肝脏阶段的寄生虫具有活性，直到寄生虫对小分子获得低水平的抗性。我将使用基因组扫描来确定可能的目标。突变将被设计到敏感的寄生虫菌株中，以证明它们会引起抗药性。将使用免疫和电子显微镜和定位研究以及破坏研究进一步表征可能的靶点。这项工作可能会导致更好地了解如何治疗组织阶段疟疾，提供新的抗生素耐药基因，并提供有关真核病原体如何对药物产生耐药性的信息。

DESCRIPTION (provided by applicant): Because blood stage infections produce most clinical manifestations of malaria, drug development has primarily focused on the development of schizonticides targeting Plasmodium falciparum, the causative agent of the most severe form of human malaria. Increased funding and a growing awareness of the problem of parasite resistance have helped to push a number of new schizonticides into the developmental pipeline and even clinical trials. On the other hand, few of these drug candidates are effective against malaria exoerythrocytic stages, and even fewer are likely to provide a radical cure for P. vivax malaria. The need for drugs which can act as a replacement for primaquine is even more urgent as malaria eradication becomes a higher priority for the world health community. In order to stimulate drug development activity against hepatic stages, more work is needed to understand hepatic stage biology and to discover targets whose activity is essential for both hepatic and blood stage development. I will use a chemical genetic approach to investigate pathways that are critically essential to both blood and hepatic parasite development and then characterize the target(s) revealed by this approach. Specifically, I will grow parasites under sub-lethal concentrations of small molecules with activity against blood and hepatic stage parasites until the parasites acquire low level resistance to the small molecules. I will use genome-scanning to determine the likely target(s). Mutations will be engineered into sensitive parasite strains to prove that they cause resistance. Likely targets will be further characterized using immuno and electron microscopy and localization studies, as well as disruption studies. The work may lead to a better understanding of how to treat tissue stage malaria, provide new antibiotic resistance genes, and provide information about how eukaryotic pathogens become resistant to drugs.