Dual artemisinin action combats resistance

摘要 广泛的长期目标是优化关键的青蒿素和喹啉疟疾药物组合，使其最大限度地 通过战胜抗药性杀死恶性疟原虫。在对目前存在的抗药性背景下 部署药物，这项工作将量化一种新的类似喹啉的血红素-青蒿素加合物的作用机制， 确定与耐药水平相关的可逆性或不可逆性的血红素晶体抑制，并探索最佳方案 与喹啉-青蒿素联合杀灭寄生虫有关的血红素晶体抑制。初步数据验证 青蒿素的另一个作用机制是基于形成丰富的血红素-青蒿素加合物， 它以不可逆转的作用抑制了血红素的结晶。外源性血红素青蒿素加合物对青蒿素的抑制作用 具有低NM IC50的抗环突变kelch13恶性疟原虫。该实验方法使用 恶性疟原虫药敏和耐药体外实验研究及协同作用 原子力显微镜时间分辨原位观察晶体生长获得的物理化学洞察力 在不同药物组合存在的情况下。 假设是双氢青蒿素的血红素加合物(DHA， 体内大多数青蒿素类药物的产物)使任何疟原虫分离株的滋养体敏感，这 击败青蒿素环期抵抗。我们还假设某些喹啉类药物和 血红素-双氢青蒿素加合物(H-DHA)对与血红素晶体抑制相关的寄生虫的杀灭作用优于 以及对可逆性/不可逆性血红素晶体的抑制程度。为了实现这些目标，我们将 追求三个具体目标：目标1.建立外源As的抑制浓度和作用机制 以及在药物敏感和耐药疟原虫中的生物活性H-DHA。目标2.确定可逆性或不可逆性 体内和体外H-DHA对血红素晶体的抑制作用。目的3.确定抗疟疾喹啉类药物的双重组合是否 和H-DHA加合物增强、削弱或漠视其伴侣对寄生虫的杀灭行动和寄生虫死亡率 血红素结晶。 这项拟议的研究将量化青蒿素加合物代谢产物对寄生虫的杀伤量 使滋养体阶段对青蒿素类药物敏感。这项工作还将提供基本的知识 关于消除青蒿素耐药性的机制、血红素晶体生长的可逆性程度，以及 基于相互作用效应的疟疾药物的最佳组合。

Summary The broad long-term goal is to optimize critical artemisinin and quinoline malaria drug combinations for maximum killing of the P. falciparum parasite by defeating resistance. In the setting of ongoing drug resistance to currently deployed drugs, this work will quantify a novel quinoline-like mechanism of action for the heme-artemisinin adducts, define reversible or irreversible heme crystal inhibition correlated to level of drug resistance, and explore optimum heme crystal inhibition related to parasite killing with quinoline-artemisinin combinations. Preliminary data validate an additional mechanism of action for the artemisinins based on formation of abundant heme-artemisinin adduct, which inhibits heme crystallization with irreversible action. Exogenous heme-artemisinin adducts inhibit artemisinin ring-resistant mutant Kelch13 P. falciparum parasites with low nM IC50s. The experimental approach employs the synergy between experimental investigations with P. falciparum drug-sensitive and resistant parasites in vitro and physicochemical insights obtained by time-resolved in situ observations of crystal growth by atomic force microscopy in the presence of different drug combinations. The hypothesis is that the inhibition of heme crystal formation by the heme adduct of dihydroartemisinin (DHA, the product of most artemisinin-class drugs in vivo) renders trophozoites of any Plasmodium isolate sensitive, which defeats the artemisinin ring-stage resistance. We also hypothesize that certain combinations of quinolines and heme-dihydroartemisinin adduct (H-DHA) are superior in killing of parasites correlated to heme crystal inhibition as well as separately to the degree of reversible/irreversible heme crystal inhibition. Towards these objectives, we will pursue three specific aims: Aim 1. Establish the inhibition concentrations and mechanism of action of exogenous as well as bio-activated H-DHA in drug sensitive and resistant Plasmodium. Aim 2. Establish reversibility or irreversibility of H-DHA heme crystal inhibition in vivo and in vitro. Aim 3. Establish if double combinations of antimalarial quinolines and H-DHA adducts enhance, weaken, or are indifferent to their partner’s action on parasite killing and the rate of hematin crystallization. This proposed research will quantify the amount of parasite killing by artemisinin adduct metabolites which renders trophozoite stages sensitive to the artemisinin drug class. The work will also inform fundamental knowledge regarding mechanisms to defeat artemisinin resistance, degree of reversibility of hematin crystal growth, and optimum combinations of malaria drugs based upon interaction effects.