Biosciences and Biotechnology Postgraduate Program, Carlos Chagas Institute, (ICC/Fiocruz/PR), Curitiba, Paraná, Brazil; State University of Londrina (UEL/PR), Laboratory of Immunoparasitology, Londrina, Paraná, Brazil.
State University of Londrina (UEL/PR), Laboratory of Biotransformation and Phytochemistry, Londrina, Paraná, Brazil.
Chem Biol Interact. 2022 Jan 5;351:109690. doi: 10.1016/j.cbi.2021.109690. Epub 2021 Oct 9.
The currently available treatment options for leishmaniasis are associated with high costs, severe side effects, and high toxicity. In previous studies, thiohydantoins demonstrated some pharmacological activities and were shown to be potential hit compounds with antileishmanial properties. The present study further explored the antileishmanial effect of acetyl-thiohydantoins against Leishmania amazonensis and determined the main processes involved in parasite death. We observed that compared to thiohydantoin nuclei, acetyl-thiohydantoin treatment inhibited the proliferation of promastigotes. This treatment caused alterations in cell cycle progression and parasite size and caused morphological and ultrastructural changes. We then investigated the mechanisms involved in the death of the protozoan; there was an increase in ROS production, phosphatidylserine exposure, and plasma membrane permeabilization and a loss of mitochondrial membrane potential, resulting in an accumulation of lipid bodies and the formation of autophagic vacuoles on these parasites and confirming an apoptosis-like process. In intracellular amastigotes, selected acetyl-thiohydantoins reduced the percentage of infected macrophages and the number of amastigotes/macrophages by increasing ROS production and reducing TNF-α levels. Moreover, thiohydantoins did not induce cytotoxicity in murine macrophages (J774A.1), human monocytes (THP-1), or sheep erythrocytes. In silico and in vitro analyses showed that acetyl-thiohydantoins exerted in vitro antileishmanial effects on L. amazonensis promastigotes in apoptosis-like and amastigote forms by inducing ROS production and reducing TNF-α levels, indicating that they are good candidates for drug discovery studies in leishmaniasis treatment. Additionally, we carried out molecular docking analyses of acetyl-thiohydantoins on two important targets of Leishmania amazonensis: arginase and TNF-alpha converting enzyme. The results suggested that the acetyl groups in the N1-position of the thiohydantoin ring and the ring itself could be pharmacophoric groups due to their affinity for binding amino acid residues at the active site of both enzymes via hydrogen bond interactions. These results demonstrate that thiohydantoins are promising hit compounds that could be used as antileishmanial agents.
目前用于治疗利什曼病的方法存在成本高、副作用严重和毒性大等问题。在以前的研究中，硫代海因衍生物表现出一些药理活性，被认为是具有抗利什曼原虫特性的潜在先导化合物。本研究进一步探索了乙酰硫代海因衍生物对亚马逊利什曼原虫的抗利什曼原虫作用，并确定了参与寄生虫死亡的主要过程。我们观察到，与硫代海因核相比，乙酰硫代海因处理抑制了前鞭毛体的增殖。这种处理导致细胞周期进程和寄生虫大小的改变，并导致形态和超微结构的变化。然后，我们研究了参与原生动物死亡的机制；活性氧（ROS）的产生增加，磷脂酰丝氨酸暴露和质膜通透性增加，线粒体膜电位丧失，导致脂滴在这些寄生虫上的积累和自噬空泡的形成，并证实了一种类似凋亡的过程。在细胞内无鞭毛体中，所选乙酰硫代海因通过增加 ROS 的产生和降低 TNF-α 水平，减少感染巨噬细胞的百分比和巨噬细胞内无鞭毛体的数量。此外，硫代海因在小鼠巨噬细胞（J774A.1）、人单核细胞（THP-1）或绵羊红细胞中没有诱导细胞毒性。在计算机和体外分析表明，乙酰硫代海因通过诱导 ROS 的产生和降低 TNF-α 水平，对亚马逊利什曼原虫的前鞭毛体形式和无鞭毛体形式发挥体外抗利什曼原虫作用，表明它们是利什曼病治疗药物发现研究的良好候选药物。此外，我们对亚马逊利什曼原虫的两个重要靶标：精氨酸酶和 TNF-α 转化酶，进行了乙酰硫代海因的分子对接分析。结果表明，硫代海因环的 N1 位上的乙酰基和环本身可能是药效团，因为它们通过氢键相互作用与两个酶的活性位点上的氨基酸残基结合的亲和力。这些结果表明，硫代海因衍生物是有前途的潜在先导化合物，可作为抗利什曼原虫药物。
