Samuel B U, Hearn B, Mack D, Wender P, Rothbard J, Kirisits M J, Mui E, Wernimont S, Roberts C W, Muench S P, Rice D W, Prigge S T, Law A B, McLeod R
Department of Visual Sciences, University of Chicago, 5841 South Maryland, AMB S-208, Chicago, IL 60637, USA.
Proc Natl Acad Sci U S A. 2003 Nov 25;100(24):14281-6. doi: 10.1073/pnas.2436169100. Epub 2003 Nov 17.
To eliminate apicomplexan parasites, inhibitory compounds must cross host cell, parasitophorous vacuole, and parasite membranes and cyst walls, making delivery challenging. Here, we show that short oligomers of arginine enter Toxoplasma gondii tachyzoites and encysted bradyzoites. Triclosan, which inhibits enoyl-ACP reductase (ENR), conjugated to arginine oligomers enters extracellular tachyzoites, host cells, tachyzoites inside parasitophorous vacuoles within host cells, extracellular bradyzoites, and bradyzoites within cysts. We identify, clone, and sequence T. gondii enr and produce and characterize enzymatically active, recombinant ENR. This enzyme has the requisite amino acids to bind triclosan. Triclosan released after conjugation to octaarginine via a readily hydrolyzable ester linkage inhibits ENR activity, tachyzoites in vitro, and tachyzoites in mice. Delivery of an inhibitor to a microorganism via conjugation to octaarginine provides an approach to transporting antimicrobials and other small molecules to sequestered parasites, a model system to characterize transport across multiple membrane barriers and structures, a widely applicable paradigm for treatment of active and encysted apicomplexan and other infections, and a generic proof of principle for a mechanism of medicine delivery.
为了消除顶复门寄生虫，抑制性化合物必须穿过宿主细胞、寄生泡和寄生虫膜以及包囊壁，这使得递送具有挑战性。在此，我们表明精氨酸短寡聚物可进入刚地弓形虫速殖子和包囊缓殖子。与精氨酸寡聚物偶联的三氯生可抑制烯酰基-ACP还原酶（ENR），它能进入细胞外速殖子、宿主细胞、宿主细胞内寄生泡中的速殖子、细胞外缓殖子以及包囊内的缓殖子。我们鉴定、克隆并测序了弓形虫的enr基因，并制备和表征了具有酶活性的重组ENR。该酶具有结合三氯生所需的氨基酸。通过易水解的酯键与八聚精氨酸偶联后释放的三氯生可抑制ENR活性、体外速殖子以及小鼠体内的速殖子。通过与八聚精氨酸偶联将抑制剂递送至微生物，为将抗菌剂和其他小分子运输至隐匿的寄生虫提供了一种方法，为表征跨多个膜屏障和结构的运输提供了一个模型系统，为治疗活跃和包囊化的顶复门寄生虫及其他感染提供了一种广泛适用的范例，以及为药物递送机制提供了一个通用的原理证明。
