Dissecting the molecular basis of PfCRT-mediated antimalarial drug resistance

 描述(申请人提供)：原虫寄生虫恶性疟原虫是人类最致命的疟疾形式，其并发症包括昏迷、贫血、呼吸窘迫和肾功能衰竭。由于手头没有有效的疫苗，化疗仍然是疟疾治疗和控制的基石。对氯喹(CQ)的抗药性上升阻碍了根除疟疾的努力，氯喹本来是一种安全、廉价和高效的药物。CQ抗性(CQR)主要由恶性疟原虫氯喹抗性转运蛋白(PfCRT)介导。除了介导CQR，PfCRT还是寄生虫对基于青蒿素的一线联合疗法(ACTS)的敏感性的调节器，其疗效因东南亚新出现的多药耐药(MDR)而受到威胁。虽然所有地理上不同的CQR寄生虫都在PfCRT中获得了多个单核苷酸多态(SNPs)，可能是为了平衡它们对耐药性和适应性的要求，但我们目前无法解释不同地区PfCRT单倍型流行的时间变化。为了了解特定的PfCRT SNP如何影响寄生虫的耐药性和适合性(目标1和2)，我们将从基因上剖析两个CQR pfcrt等位基因，厄瓜多尔1110(来自厄瓜多尔)和Cam734(来自柬埔寨)。厄瓜多尔1110 pfcrt含有4个SNPs，是观察到的最少的，这使得它适合于对导致获得耐药性的突变轨迹的研究。Cam734 pfcrt是迄今发现的突变程度最高的等位基因，它含有5个新的SNPs，为研究pfcrt提供的寄生虫适合度的独特获得提供了一个原型。我们还将 探索PfCRT SNPs促进对一线ACT成分(目标3)，即青蒿素(ARTS)及其伙伴药物的耐药性的能力，这一努力将结合我们基于实验室的研究与柬埔寨西部的现场临床培训，柬埔寨西部是MDR的震中。综上所述，我们的目标是(1)调查PfCRT SNPs在CQ和其他临床上重要的抗疟疾药物耐药形成中的作用，(2)评估 PfCRT SNPs对寄生虫适合性的影响，以及(3)评估PfCRT调节多药耐药寄生虫对ARTS和ACT伙伴药物敏感性的能力。这些研究有望对抗疟疾耐药性的遗传基础产生重要的新见解，包括pfcrt突变在调节一线联合治疗疗效中的作用，并将指导开发新的战略，以减少多药耐药疟疾的全球影响。

 DESCRIPTION (provided by applicant): The protozoan parasite Plasmodium falciparum is responsible for the deadliest form of human malaria, with complications including coma, anemia, respiratory distress, and renal failure. With no effective vaccine at hand, chemotherapy remains the cornerstone for malaria treatment and control. Efforts to eradicate malaria have been hindered by the rise of resistance to chloroquine (CQ), an otherwise safe, cheap, and highly effective drug. CQ resistance (CQR) is primarily mediated by the P. falciparum Chloroquine Resistance Transporter (PfCRT). In addition to mediating CQR, PfCRT is a modulator of parasite susceptibility to first-line artemisinin- based combination therapies (ACTs), whose efficacy is endangered by emerging multidrug resistance (MDR) in Southeast Asia. While all geographically-distinct CQR parasites have acquired multiple single-nucleotide polymorphisms (SNPs) in PfCRT, presumably to balance their requirements for drug resistance and fitness, we are currently unable to explain temporal changes in the prevalence of different regional PfCRT haplotypes. To understand how specific PfCRT SNPs affect parasite drug resistance and fitness (Aims 1 and 2), we will genetically dissect two CQR pfcrt alleles, Ecu1110 (from Ecuador) and Cam734 (from Cambodia). Ecu1110 pfcrt contains 4 SNPs, the fewest observed, rendering it amenable to a study of the mutational trajectories that lead to acquisition of drug resistance. Cam734 pfcrt, the most highly mutated allele yet described, harbors 5 novel SNPs and serves as a prototype for the study of the unique gain of parasite fitness afforded by pfcrt. We will also explore the capacity of PfCRT SNPs to facilitate resistance to first-line ACT components (Aim 3), namely artemisinins (ARTs) and their partner drugs, an effort that will combine our laboratory-based investigations with on-site clinical training in western Cambodia, an epicenter of MDR. In summary, our aims are (1) to investigate the contribution of PfCRT SNPs to the development of resistance to CQ and other clinically significant antimalarials, (2) to evaluate the impact of PfCRT SNPs on parasite fitness, and (3) to assess the capacity of PfCRT to modulate susceptibility to ARTs and ACT partner drugs in multidrug-resistant parasites. These studies are expected to yield important new insights into the genetic basis of antimalarial drug resistance, including the role of pfcrt mutations in modulating the efficacy of first-line combination therapie, and will guide the development of novel strategies to reduce the global impact of multidrug-resistant malaria.