Modeling mutant PfCRT-mediated drug transport to predict the emergence of piperaquine-resistant Plasmodium falciparum malaria

模拟突变 PfCRT 介导的药物转运以预测耐哌喹恶性疟原虫疟疾的出现

基本信息

批准号：
10326368
负责人：
Laura Marie Hagenah
金额：
$ 4.68万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-01-01 至 2023-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10326368
关键词：
Affect Africa African African American Alleles Amino Acid Substitution Amodiaquine Anti-malarial drug resistance Antimalarials Area Asia Asian Binding Biological Assay Biophysics Blood Cessation of life Charge Child Chloroquine Chloroquine resistance Clinical Coculture Techniques Combined Modality Therapy Cryoelectron Microscopy Data Drug Efflux Drug Transport Drug resistance Engineering Exhibits Falciparum Malaria Flow Cytometry Genes Genetic Genetic Engineering Geographic Locations Geography Globin Goals Growth Haplotypes Hemoglobin In Vitro Kinetics Knowledge Malaria Malaria Vaccines Mediating Methods Modeling Molecular Morbidity - disease rate Mutation Organelles Parasites Peptide Transport Peptides Persons Pharmaceutical Preparations Physiological Plasmodium falciparum Predisposition Property Protein Biosynthesis Protein Isoforms Recombinant Proteins Regimen Reporter Research Resistance Resolution Risk Role Site South America South American Southeastern Asia Structure Susceptibility Gene System Techniques Testing Therapeutic Time Treatment Protocols Universities Vaccines Vacuole Variant asexual base chemotherapy combat cost disorder control fitness global health insight mortality mutant new combination therapies nonsynonymous mutation novel resistance mutation resistant Plasmodium falciparum resistant strain socioeconomics therapy design therapy development treatment strategy

项目摘要

PROJECT SUMMARY The protozoan parasite Plasmodium falciparum causes over 400,000 malaria deaths each year, mostly in young African children. With no effective vaccine, chemotherapy remains the cornerstone of malaria treatment and control. Malaria eradication efforts have been hindered by the rise of resistance to first-line antimalarials in Southeast Asia, including piperaquine (PPQ, used in combination with dihydroartemisinin). Piperaquine resistance (PPQ-R) and chloroquine resistance are primarily mediated by mutations in the P. falciparum Chloroquine Resistance Transporter (PfCRT). Mutant PfCRT transports drug away from its site of action in the asexual blood stage parasite’s digestive vacuole. This acidic organelle degrades endocytosed host hemoglobin and extrudes globin-derived peptides for parasite protein synthesis. Less understood, however, is how certain amino acid substitutions confer this efflux mechanism and how they impact native transporter function. To investigate the effect of these mutations, I propose in Aim 1 to purify contemporary PPQ-R PfCRT isoforms and perform binding and transport assays with PPQ, other clinically used antimalarials, and positively-charged peptides as proposed natural substrates. These studies will provide a comprehensive functional characterization of PPQ-R PfCRT that is currently lacking. It is also important to predict how PPQ-R could spread to or emerge in other malaria endemic regions. The PfCRT mutations found in SE Asia have yet to be seen on African and South American backgrounds; however, PPQ is being used in these areas including as first-line treatment or for uncomplicated malaria. To predict whether the contemporary amino acid substitutions seen in SE Asia could emerge in other regions to achieve PPQ-R, I propose in Aim 2 to engineer these mutations onto African and South American PfCRT isoforms in P. falciparum parasites from these regions. Assays will quantify the susceptibility of these lines to PPQ and other antimalarials, and determine the relative fitness of each line. The degree of resistance conferred and the fitness cost imposed, along with the regional drug regimen, will be important in determining which pfcrt alleles predominate and which can emerge and spread in areas of PPQ use. These aims are predicated on the hypotheses that (1) PPQ-R mutations in PfCRT alter transport of drugs and natural substrates and that (2) PPQ-R can arise in Africa and South America via the emergence of single amino acid substitutions observed in SE Asia in mutant PfCRT. These data will also identify whether a gain of PPQ-R restores parasite susceptibility to chloroquine, as has been seen with most mutations in SE Asia, thereby creating therapeutic opportunities for new combination therapies. These studies are expected to yield important new insights into the molecular basis for antimalarial drug resistance, and to leverage that knowledge to predict the emergence of novel PPQ-R PfCRT isoforms in distinct geographic regions. This will guide the development of treatment strategies to reduce the global impact and spread of drug-resistant malaria.

项目概要原生动物寄生虫恶性疟原虫每年导致超过 400,000 人死于疟疾，其中大多数是年轻人由于没有有效的疫苗，化疗仍然是非洲儿童治疗疟疾的基石。由于一线抗疟药耐药性的增加，根除疟疾的努力受到阻碍。东南亚，包括哌喹（PPQ，与二氢青蒿素联合使用）。耐药性（PPQ-R）和氯喹耐药性主要是由恶性疟原虫的突变介导的氯喹抗性转运蛋白 (PfCRT) 突变体 PfCRT 将药物转运离开其作用位点。无性血液期寄生虫的消化液泡这种酸性细胞器会降解内吞的宿主血红蛋白。并挤出球蛋白衍生的肽用于寄生虫蛋白质的合成，然而，其确定性却鲜为人知。氨基酸取代赋予了这种外排机制以及它们如何影响天然转运蛋白功能。为了研究这些突变的影响，我在目标 1 中建议纯化当代 PPQ-R PfCRT 同工型，使用 PPQ、其他临床使用的抗疟药和带正电的药物进行结合和转运测定这些研究将提供全面的功能表征。预测 PPQ-R 如何传播或出现也很重要。在其他疟疾流行地区，尚未在非洲和非洲发现 PfCRT 突变。南美背景；然而，PPQ 正在这些领域使用，包括作为一线治疗或用于预测东南亚的现代氨基酸替代是否可以治疗无并发症的疟疾。为了实现 PPQ-R，我在目标 2 中建议将这些突变设计到非洲和南美恶性疟原虫中的 PfCRT 同种型检测将量化这些地区的恶性疟原虫寄生虫。这些品系对 PPQ 和其他抗疟药的敏感性，并确定每个品系的相对适合度。赋予的抵抗程度和施加的健身成本以及区域药物治疗方案将对于确定哪些 pfcrt 等位基因占主导地位以及哪些可以在 PPQ 区域出现和传播很重要这些目标基于以下假设：(1) PfCRT 中的 PPQ-R 突变改变药物转运。和天然底物，并且 (2) PPQ-R 可以通过单一物质的出现在非洲和南美洲出现在东南亚观察到的突变体 PfCRT 中的氨基酸取代这些数据也将确定是否有增益。 PPQ-R 恢复了寄生虫对氯喹的敏感性，正如在东南亚的大多数突变中所看到的那样，从而这些研究预计将产生重要成果。对抗疟药物耐药性的分子基础的新见解，并利用这些知识来预测新型 PPQ-R PfCRT 同种型在不同地理区域的出现将指导开发。减少耐药性疟疾的全球影响和传播的治疗策略。