Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892.
Center for Global Health and Infectious Diseases Research, College of Public Health, University of South Florida, Tampa, FL 33612.
Proc Natl Acad Sci U S A. 2018 Dec 4;115(49):12513-12518. doi: 10.1073/pnas.1813386115. Epub 2018 Nov 19.
Concerns about malaria parasite resistance to treatment with artemisinin drugs (ARTs) have grown with findings of prolonged parasite clearance s (>5 h) and their association with mutations in Kelch-propeller protein K13. Here, we describe a laboratory cross of K13 C580Y mutant with C580 wild-type parasites to investigate ART response phenotypes in vitro and in vivo. After genotyping >400 isolated progeny, we evaluated 20 recombinants in vitro: IC measurements of dihydroartemisinin were at similar low nanomolar levels for C580Y- and C580-type progeny (mean ratio, 1.00; 95% CI, 0.62-1.61), whereas, in a ring-stage survival assay, the C580Y-type progeny had 19.6-fold (95% CI, 9.76-39.2) higher average counts. In splenectomized monkeys treated with three daily doses of i.v. artesunate, calculations by three different methods yielded mean differences of 0.01 h (95% CI, -3.66 to 3.67), 0.80 h (95% CI, -0.92 to 2.53), and 2.07 h (95% CI, 0.77-3.36) between C580Y and C580 infections. Incidences of recrudescence were 57% in C580Y (4 of 7) versus 70% in C580 (7 of 10) infections (-13% difference; 95% CI, -58% to 35%). Allelic substitution of C580 in a C580Y-containing progeny clone (76H10) yielded a transformant (76H10) that, in an infected monkey, recrudesced regularly 13 times over 500 d. Frequent recrudescences of ART-treated infections occur with or without K13 mutations and emphasize the need for improved partner drugs to effectively eliminate the parasites that persist through the ART component of combination therapy.
人们对疟原虫对青蒿素类药物（ARTs）治疗的耐药性越来越担忧，因为发现寄生虫清除时间延长（>5 小时），并且与 Kelch-propeller 蛋白 K13 中的突变有关。在这里，我们描述了 K13 C580Y 突变体与 C580 野生型寄生虫的实验室杂交，以研究体外和体内的 ART 反应表型。在对>400 个分离后代进行基因分型后，我们评估了 20 个重组体：二氢青蒿素的 IC 测量值对于 C580Y-和 C580 型后代相似，处于低纳摩尔水平（平均值比，1.00；95%CI，0.62-1.61），而在环状阶段生存测定中，C580Y 型后代的平均计数高 19.6 倍（95%CI，9.76-39.2）。在接受静脉注射青蒿琥酯每日 3 次治疗的脾切除猴子中，通过三种不同方法计算得出的平均差异为 0.01 小时（95%CI，-3.66 至 3.67），0.80 小时（95%CI，-0.92 至 2.53）和 2.07 小时（95%CI，0.77-3.36），C580Y 和 C580 感染之间。C580Y 感染的复发率为 57%（4/7），而 C580 感染的复发率为 70%（7/10）（-13%差异；95%CI，-58%至 35%）。在含有 C580Y 的后代克隆（76H10）中，C580 的等位基因替代产生了一个转化体（76H10），在感染猴子中，在 500 天内定期复发 13 次。ART 治疗的感染经常出现复发，无论是否存在 K13 突变，这强调了需要更好的联合药物来有效消除通过联合治疗中的 ART 成分持续存在的寄生虫。
