Reyes-Vivas Horacio, de la Mora-de la Mora Ignacio, Castillo-Villanueva Adriana, Yépez-Mulia Lilian, Hernández-Alcántara Gloria, Figueroa-Salazar Rosalia, García-Torres Itzhel, Gómez-Manzo Saúl, Méndez Sara T, Vanoye-Carlo América, Marcial-Quino Jaime, Torres-Arroyo Angélica, Oria-Hernández Jesús, Gutiérrez-Castrellón Pedro, Enríquez-Flores Sergio, López-Velázquez Gabriel
Laboratorio de Bioquímica Genética, Instituto Nacional de Pediatría, Mexico City, Mexico.
Centro Médico Nacional Siglo XXI, IMSS, Mexico City, Mexico.
Antimicrob Agents Chemother. 2014 Dec;58(12):7072-82. doi: 10.1128/AAC.02900-14. Epub 2014 Sep 15.
Giardiasis is highly prevalent in the developing world, and treatment failures with the standard drugs are common. This work deals with the proposal of omeprazole as a novel antigiardial drug, focusing on a giardial glycolytic enzyme used to follow the cytotoxic effect at the molecular level. We used recombinant technology and enzyme inactivation to demonstrate the capacity of omeprazole to inactivate giardial triosephosphate isomerase, with no adverse effects on its human counterpart. To establish the specific target in the enzyme, we used single mutants of every cysteine residue in triosephosphate isomerase. The effect on cellular triosephosphate isomerase was evaluated by following the remnant enzyme activity on trophozoites treated with omeprazole. The interaction of omeprazole with giardial proteins was analyzed by fluorescence spectroscopy. The susceptibility to omeprazole of drug-susceptible and drug-resistant strains of Giardia lamblia was evaluated to demonstrate its potential as a novel antigiardial drug. Our results demonstrate that omeprazole inhibits giardial triosephosphate isomerase in a species-specific manner through interaction with cysteine at position 222. Omeprazole enters the cytoplasmic compartment of the trophozoites and inhibits cellular triosephosphate isomerase activity in a dose-dependent manner. Such inhibition takes place concomitantly with the cytotoxic effect caused by omeprazole on trophozoites. G. lamblia triosephosphate isomerase (GlTIM) is a cytoplasmic protein which can help analyses of how omeprazole works against the proteins of this parasite and in the effort to understand its mechanism of cytotoxicity. Our results demonstrate the mechanism of giardial triosephosphate isomerase inhibition by omeprazole and show that this drug is effective in vitro against drug-resistant and drug-susceptible strains of G. lamblia.
贾第虫病在发展中世界高度流行，使用标准药物治疗失败的情况很常见。这项工作涉及将奥美拉唑作为一种新型抗贾第虫药物的提议，重点关注一种用于在分子水平追踪细胞毒性作用的贾第虫糖酵解酶。我们使用重组技术和酶失活来证明奥美拉唑使贾第虫磷酸丙糖异构酶失活的能力，而对其人类对应物没有不良影响。为了确定该酶中的特定靶点，我们使用了磷酸丙糖异构酶中每个半胱氨酸残基的单突变体。通过追踪用奥美拉唑处理的滋养体上的残余酶活性来评估对细胞磷酸丙糖异构酶的影响。通过荧光光谱分析奥美拉唑与贾第虫蛋白质的相互作用。评估了药物敏感和耐药的蓝氏贾第鞭毛虫菌株对奥美拉唑的敏感性，以证明其作为新型抗贾第虫药物的潜力。我们的结果表明，奥美拉唑通过与222位的半胱氨酸相互作用，以物种特异性方式抑制贾第虫磷酸丙糖异构酶。奥美拉唑进入滋养体的细胞质区室，并以剂量依赖性方式抑制细胞磷酸丙糖异构酶活性。这种抑制与奥美拉唑对滋养体引起的细胞毒性作用同时发生。蓝氏贾第鞭毛虫磷酸丙糖异构酶（GlTIM）是一种细胞质蛋白，有助于分析奥美拉唑如何作用于这种寄生虫的蛋白质，并有助于理解其细胞毒性机制。我们的结果证明了奥美拉唑抑制贾第虫磷酸丙糖异构酶的机制，并表明该药物在体外对蓝氏贾第鞭毛虫的耐药和药物敏感菌株均有效。
