Molecular Pathways Affected by Drugs that Disrupt Na+ Homeostasis in Malaria Parasites

每年有数亿疟疾病例接受抗疟药物治疗， 个别病人携带数千亿疟疾寄生虫，有必要继续喂养 新的抗疟药物管道，以应对可能出现的耐药性。近年来，一些 新的抗疟化合物已经被发现具有破坏疟疾中Na+稳态的能力 寄生虫其中四种（螺吲哚酮、吡唑酰胺、二氢异喹诺酮和硫代三唑）具有 被指定为临床候选药物。值得注意的是，这些药物属于非常不同的化学类别 具有不同的药效团和针对疟原虫生命周期不同阶段的活性。重要的是所有 这些药物显示出体内寄生虫的快速清除。对这些化合物中的几种具有抗性的寄生虫 显示了P型ATP酶PfATP 4内的一系列突变，现在认为PfATP 4是Na+泵。因此，在本发明中， 通过抑制PfATP 4的Na+内流被认为是所有这些的共同作用机制。 化合物.我们在过去几年的研究表明，虽然PfATP 4的突变是必要的， 尽管对所有这些化合物都有抗性，但它们并不总是足以产生完全水平的抗性。我们 他们发现，抗吡唑酰胺的寄生虫携带额外的突变，这是赋予完整的基因所必需的。 与PfATP 4突变相关的耐药性，表明PfATP 4的上位调控成分 活动我们对Na+流入寄生虫的生理后果的研究揭示了一些 戏剧性的变化表明一个迄今未知的调节途径，被不适当的干扰， 寄生虫胞质Na+水平。因此，彻底调查受影响的分子途径 疟疾寄生虫中Na+稳态的破坏既有必要，也有可能提供进一步的见解， 指导未来的药物发现和开发。基因编辑和条件生物学技术的最新进展 恶性疟原虫的基因表达使我们有可能以前所未有的方式解开这些途径。 续费通过应用这些方法，我们将研究PfATP 4在维持Na+和Na +-ATP酶中的作用。 恶性疟原虫胆固醇稳态。我们将评估耐药相关的表型后果， PfATP 4突变，并研究PfATP 4以外的基因突变在影响耐药性中的作用。 与PfATP 4突变的组合。我们将研究一个假定的质膜胆固醇转运蛋白 这些新的抗疟疾药物的影响。这些研究将促进我们对新分子的理解， 我们已经验证了这些途径，作为正在开发的有效抗疟疾药物的靶点。

With hundreds of millions of malaria cases being treated with antimalarial drugs each year and with each individual patient bearing hundreds of billions of malaria parasites, it is necessary to continue to feed the antimalarial pipeline with new drugs to counter the likely emergence of resistance. In recent years several novel antimalarial compounds have been discovered with the ability to disrupt Na+ homeostasis in malaria parasites. Four of these (a spiroindolone, a pyrazoleamide, a dihydroisoquinolone, and a thiotriazole) have been designated clinical drug candidates. Remarkably, these drugs belong to very different chemical classes with distinct pharmacophores and activity against different stages of malaria parasite life cycle. Importantly, all these drugs show fast clearance of parasites in vivo. Parasites resistant to several of these compounds have shown a range of mutations within a P-type ATPase, PfATP4, that is now believed to be a Na+ pump. Thus, influx of Na+ through inhibition of PfATP4 is considered to be the common mechanism of action for all these compounds. Our work over the last few years has revealed that, while mutations in PfATP4 are necessary for resistance to all of these compounds, they are not always sufficient to generate the full level of resistance. We have found that pyrazoleamide-resistant parasites bear additional mutations, which are required to impart full resistance in conjunction with PfATP4 mutations, suggesting epistatic regulatory components to PfATP4 activity. Our investigations of physiological consequences of Na+ influx into the parasite have revealed some dramatic changes suggesting a hitherto unknown regulatory pathway that is perturbed by inappropriate cytosolic Na+ levels in the parasite. Therefore, a thorough investigation of molecular pathways affected by disruption of Na+ homeostasis in malaria parasites is both necessary and likely to provide further insights to guide future drug discovery and development. Recent advances in technology for gene editing and conditional gene expression in Plasmodium falciparum make it now possible to unravel these pathways in unprecedented details. By applying these approaches, we will investigate the role of PfATP4 in maintenance of Na+ and cholesterol homeostasis in P. falciparum. We will assess phenotypic consequences of resistance-associated mutations in PfATP4, and study the role of mutations in genes other than PfATP4 that affect drug resistance in combination with PfATP4 mutations. We will investigate a putative plasma membrane cholesterol transporter that is affected by these new antimalarials. These studies will advance our understanding of novel molecular pathways that we have validated as targets for potent antimalarial drugs in development.