PNAS：反义核酸新型疗法治疗弓形体病

近日，来自芝加哥大学的研究者成功开发出了一种治疗弓形体病的新的治疗方法，弓形体病是一种寄生虫病。这种新型方法在早期试管和动物实验中表明，其可以阻止寄生虫产生选择性的蛋白质，当这种新型疗法在小鼠中进行试验的时候，研究者揭示了其可以明显降低小鼠体内超过90%的可见寄生虫。相关研究成果刊登在了近日的国际杂志PNAS上。

这种新型疗法结合了短链的反义核酸，反义核酸是一种携带小型肽类的物质，可以通过细胞膜将其它的反义核酸转运到寄生虫体内，然后破坏寄生虫体内的遗传信号转导。研究者Rima博士表示，我们使用这种新型技术可以跨过多层膜，将这种反义核酸注射入寄生虫的活细胞中，并且阻止其制造各种毒性蛋白质。因此这样一来我们就可以有效关闭寄生虫体内任何基因的表达。

不光如此，这种新型疗法在非寄生虫疾病中也可以起到作用，其可以用于治疗细菌或者病毒感染引发的疾病，包括肌肉营养不良症等。

鼠弓形虫是世界范围内常见的主要寄生虫感染疾病，1/3的人类都被这种寄生虫感染过，其可以在那些免疫系统未成熟人们体内进行感染，并且引发疾病，尤其是感染子宫中正在发育的胎儿。因此对于治疗鼠弓形虫的感染的新型疗法迫在眉睫，标准的治疗方法常常会引发副作用，而且病人会对标准疗法产生高度过敏反应。而且目前也没有有效的药物和疫苗来有效保护机体不受寄生虫的感染。

研究者开发出的新型疗法包括一种磷酸化酰胺化物吗啉低聚物（phosphorodiamidate morpholine oligomer，POM），这是一种类似于DNA的短链分子，其可以结合到信使RNA上，抑制其翻译成为蛋白质。PMO可以连接至转换式的肽类上面，随后被转换式肽跨越细胞屏障来进行运输，这种结合方式称为PPMO，

研究者在小鼠组织活细胞中检测了这种系统的效果，结果表明，其可以破坏好几种特异性蛋白质的产生。

研究者将插入含有绿色荧光蛋白和荧光素酶的基因转化到了寄生虫细胞中，检测了PPMO对可探测生物标记物的作用，然后他们将被寄生虫感染的细胞暴露于低水平的PPMO中，结果表明，寄生虫体内的生物发光和黄色荧光降低了40%-60%，也就揭示了PPMO可以明显抑制寄生虫的功能发挥。

编译自：Antisense therapy shows promise in treating toxoplasmosis

doi:10.1073/pnas.1208775109
PMC:
PMID:

Molecular target validation, antimicrobial delivery, and potential treatment of Toxoplasma gondii infections

Bo-Shiun Lai, William H. Witola1, Kamal El Bissati, Ying Zhou, Ernest Mui, Alina Fomovska, and Rima McLeod2

Toxoplasma gondii persistently infects over two billion people worldwide. It can cause substantial morbidity and mortality. Existing treatments have associated toxicities and hypersensitivity and do not eliminate encysted bradyzoites that recrudesce. New, improved medicines are needed. Transductive peptides carry small molecule cargos across multiple membranes to enter intracellular tachyzoites and encysted bradyzoites. They also carry cargos into retina when applied topically to eyes, and cross blood brain barrier when administered intravenously. Phosphorodiamidate morpholino oligomers (PMO) inhibit gene expression in a sequence-specific manner. Herein, effect of transductive peptide conjugated PMO (PPMO) on tachyzoite protein expression and replication in vitro and in vivo was studied. Initially, sequence-specific PPMO successfully reduced transfected T. gondii’s fluorescence and luminescence. PPMO directed against T. gondii’s dihydrofolate reductase (DHFR), an enzyme necessary for folate synthesis, limited tachyzoite replication. Rescue with exogenous folate demonstrated DHFR PPMO‘s specificity. PPMO directed against enoyl-ACP reductase (ENR), an enzyme of type II fatty acid synthesis that is structurally distinct in T. gondii from ENR in mammalian cells was investigated. PPMO directed against plant-like Apetela 2 (AP2) domain transcription factor XI-3 (AP2XI-3), not present in human cells, was characterized. ENR and AP2XI-3 PPMO each restricted intracellular parasite replication validating these molecular targets in tachyzoites. DHFR-specific PPMO administered to infected mice diminished parasite burden. Thus, these antisense oligomers are a versatile approach to validate T. gondii molecular targets, reduce essential T. gondii proteins in vitro and in vivo, and have potential for development as curative medicines.