Grecco Simone S, Costa-Silva Thais A, Jerz Gerold, de Sousa Fernanda S, Alves Conserva Geanne A, Mesquita Juliana T, Galuppo Mariana K, Tempone Andre G, Neves Bruno J, Andrade Carolina H, Cunha Rodrigo L O R, Uemi Miriam, Sartorelli Patricia, Lago João Henrique G
Center of Natural Sciences and Humanities, Federal University of ABC, Santo Andre, São Paulo 09210-180, Brazil; Institute of Food Chemistry, Technische Universität Braunschweig, Braunschweig, 38106, Germany.
Center of Parasitology and Mycology, Adolfo Lutz Institute, São Paulo 01246-902, Brazil.
Phytomedicine. 2017 Jan 15;24:62-67. doi: 10.1016/j.phymed.2016.11.015. Epub 2016 Nov 21.
From a previous screening of Brazilian biodiversity for antiprotozoal activity, the hexane extract from leaves of Nectandra leucantha (Nees & Mart.) (Lauraceae) demonstrated activity against Trypanosoma cruzi. Chromatographic separation of this extract afforded bioactive dehydrodieugenol (1). Furthermore, methylated derivative 2 (dehydrodieugenol dimethyl ether) was prepared and also tested against T. cruzi.
To examine the therapeutical potential of compounds 1 and 2 against T. cruzi as well as to elucidate the mechanism of action of bioactive compound 1 against T. cruzi.
METHODS/STUDY DESIGN: Crude hexane extract from leaves was subjected to chromatographic steps to afford bioactive compound 1. In order to analyze the effect of additional methyl group in the antiparasitic activity of 1, derivative 2 was prepared (both are no pan-assay interference compounds - PAINS). These compounds were evaluated in vitro against T. cruzi (trypomastigote and amastigote forms) and analyzed for the potential effect in host cells through the production of nitric oxide and reactive oxygen species. Finally, the plasma membrane effect of the most potent compound 1 was investigated in T. cruzi trypomastigotes.
Compounds 1 and 2 displayed activity against amastigotes of T. cruzi. Although both compounds promoted activity against intracellular amastigotes, the production of nitric oxide and reactive oxygen species of host cells were unaltered, suggesting an antiparasitic activity other than host cell activation. Considering 1 the most effective compound against T. cruzi, the interference in the plasma membrane of the trypomastigotes was investigated using the fluorescent probe SYTOX Green. After a short-term incubation, the fluidity and integrity of the plasma membrane was completely altered, suggesting it as a primary target for compound 1 in T. cruzi.
Compounds 1 and 2 selectively eliminated the intracellular parasites without host cell activation and could be important scaffolds for the search of new hit compounds.
通过先前对巴西生物多样性进行的抗原生动物活性筛选，白楠（樟科）叶的己烷提取物表现出对克氏锥虫的活性。该提取物经色谱分离得到具有生物活性的脱氢二丁香酚（1）。此外，制备了甲基化衍生物2（脱氢二丁香酚二甲醚）并对其进行了抗克氏锥虫测试。
研究化合物1和2对克氏锥虫的治疗潜力，并阐明生物活性化合物1对克氏锥虫的作用机制。
方法/研究设计：叶的粗己烷提取物经色谱步骤得到生物活性化合物1。为了分析额外甲基对1的抗寄生虫活性的影响，制备了衍生物2（两者均为无泛测定干扰化合物 - PAINS）。这些化合物在体外针对克氏锥虫（锥鞭毛体和无鞭毛体形式）进行评估，并通过一氧化氮和活性氧的产生分析其对宿主细胞的潜在影响。最后，在克氏锥虫锥鞭毛体中研究了最有效的化合物1对质膜的影响。
化合物1和2对克氏锥虫无鞭毛体具有活性。尽管两种化合物均对细胞内无鞭毛体有活性，但宿主细胞的一氧化氮和活性氧产生未改变，提示其具有除宿主细胞激活以外的抗寄生虫活性。鉴于1是针对克氏锥虫最有效的化合物，使用荧光探针SYTOX Green研究了其对锥鞭毛体质膜的干扰。短期孵育后，质膜的流动性和完整性完全改变，表明其是化合物1在克氏锥虫中的主要靶点。
化合物1和2在不激活宿主细胞的情况下选择性地消除细胞内寄生虫，可能是寻找新的先导化合物的重要骨架。
