National Center for Advancing Translational Sciences, National Institutes of Health, Bethesda, MD, USA.
Nuffield Department of Medicine, University of Oxford, Oxford, UK.
Malar J. 2018 Oct 26;17(1):391. doi: 10.1186/s12936-018-2532-x.
Artemisinin-resistant Plasmodium falciparum has been reported throughout the Greater Mekong subregion and threatens to disrupt current malaria control efforts worldwide. Polymorphisms in kelch13 have been associated with clinical and in vitro resistance phenotypes; however, several studies suggest that the genetic determinants of resistance may involve multiple genes. Current proposed mechanisms of resistance conferred by polymorphisms in kelch13 hint at a connection to an autophagy-like pathway in P. falciparum.
A SNP in autophagy-related gene 18 (atg18) was associated with long parasite clearance half-life in patients following artemisinin-based combination therapy. This gene encodes PfAtg18, which is shown to be similar to the mammalian/yeast homologue WIPI/Atg18 in terms of structure, binding abilities, and ability to form puncta in response to stress. To investigate the contribution of this polymorphism, the atg18 gene was edited using CRISPR/Cas9 to introduce a T38I mutation into a k13-edited Dd2 parasite. The presence of this SNP confers a fitness advantage by enabling parasites to grow faster in nutrient-limited settings. The mutant and parent parasites were screened against drug libraries of 6349 unique compounds. While the SNP did not modulate the parasite's susceptibility to any of the anti-malarial compounds using a 72-h drug pulse, it did alter the parasite's susceptibility to 227 other compounds.
These results suggest that the atg18 T38I polymorphism may provide additional resistance against artemisinin derivatives, but not partner drugs, even in the absence of kelch13 mutations, and may also be important in parasite survival during nutrient deprivation.
青蒿素耐药恶性疟原虫已在大湄公河次区域各地出现，并有可能破坏全球当前的疟疾控制工作。kelch13 中的多态性与临床和体外耐药表型相关；然而，多项研究表明，耐药的遗传决定因素可能涉及多个基因。kelch13 多态性赋予的耐药机制目前提出的机制暗示与恶性疟原虫中的自噬样途径有关。
在接受基于青蒿素的联合疗法后，患者寄生虫清除半衰期较长与自噬相关基因 18（atg18）中的 SNP 相关。该基因编码 PfAtg18，其结构、结合能力和对应激的形成点状结构的能力与哺乳动物/酵母同源物 WIPI/Atg18 相似。为了研究该多态性的贡献，使用 CRISPR/Cas9 编辑 atg18 基因，在 k13 编辑的 Dd2 寄生虫中引入 T38I 突变。该 SNP 的存在通过使寄生虫在营养有限的环境中更快地生长而赋予其适应优势。对突变体和亲本寄生虫进行了针对 6349 种独特化合物药物库的筛选。虽然该 SNP 在用 72 小时药物脉冲处理时不会改变寄生虫对任何抗疟化合物的敏感性，但它确实改变了寄生虫对 227 种其他化合物的敏感性。
这些结果表明，atg18 T38I 多态性即使在缺乏 kelch13 突变的情况下，也可能为青蒿素衍生物提供额外的耐药性，但不是伴侣药物，并且在营养剥夺期间寄生虫的生存中也可能很重要。
