Discovery of chemically validated malaria liver stage targets

 描述(由申请人提供)：咪唑并哌嗪(IZP)是临床上用于治疗简单疟疾和疟疾预防的一类新的抗疟疾药物。IZP消除了血液阶段感染，并在疟疾动物模型中防止了无症状疟疾的发展和疟疾的传播。这一类中的一名成员已在早期临床试验中进行了测试，在口服时人类耐受性良好。虽然恶性疟原虫环胺抗性基因(PfCARL)编码突变是恶性疟原虫对IZP的抗性，但IZP的作用机制尚不清楚。因此，需要进一步研究IZP是如何发挥作用的，我们建议使用一套基因组学方法，包括代谢组学、转录组学和其他全基因组方法来研究IZP如何作用于寄生于人类的所有疟原虫生命周期形式，目的是确定它们的确切靶点。具体地说，我们将首先使用代谢组学分析来检查寄生虫代谢途径如何在血液阶段对寄生虫IZP治疗做出反应。然后，我们将比较这些对IZP的代谢反应与IZP治疗在血液期、肝期和配子体中的转录反应。我们还将对已知作用机制的化合物进行类似的分析，以确定IZP的特定作用。其次，我们将使用成熟的体外进化实验管道探索任何额外的靶点，然后进行全基因组测序。发现耐药突变的基因将与代谢和转录图谱确定的途径相关联，以确定假定的靶点。最后，我们将通过一系列的功能研究来确定恶性疟原虫中任何可能的靶点。

 DESCRIPTION (provided by applicant): Imidazolopiperazines (IZPs) are a novel class of antimalarial drug in clinical development for the treatment of uncomplicated malaria and malaria prophylaxis. IZPs eliminate blood stage infections and both prevent asymptomatic malaria development and malaria transmission in animal models of malaria. One member of the class has been tested in early clinical trials and is well-tolerated by humans when administered orally. Although parasite resistance to IZPs is conferred by mutations in the Plasmodium falciparum gene encoding the parasite P. falciparum cyclic amine resistance locus (PfCARL), the mechanism of action of the IZP's is not well understood. Further studies on how the IZPs work are therefore needed, We propose to use a suite of genomic methods including metabolomics, transcriptomics and other genome-wide methods to study how IZPs act against all Plasmodium lifecycle forms that parasitize humans, with the goal of identifying their precise target. Specifically, we will first use metabolomics profiling to examine how parasite metabolic pathways respond to parasite IZP treatment in blood stages. We will then compare these metabolic responses to the IZPs to the transcriptional response to IZP treatment in blood stages, liver stages and gametocytes. We will also perform similar analyses against compounds with known mechanisms of action, in order to identify IZP specific effects. Second, we will probe for any additional targets using a well-established experimental pipeline of in vitro evolution followed by whole genome sequencing. Genes with identified resistance mutations will be associated with the pathways identified by metabolomic and transcriptional profiling to identify putative targets. Finally, we will confirm any putative targets in P. falciparum through a series o functional studies.