HIV蛋白酶抑制剂类药物抗多房棘球蚴效果及机制研究

Echinococcosis (Hydatid disease) has a great impact on the public health and livestock husbandries in Western China. However, chemotherapeutical treatments against echinococcosis have only relied on the benzimidazoles, which only exert parasitostatic effects. There is a clear demand for new alternative drug treatment options. We have identified a functionally important gene that encodes DNA-damage inducible protein 1 (Ddi1) in the cestode genome through comparative genomic analyses. This Ddi1 protein shares a highly similar 3D structure with HIV, suggesting that the HIV protease inhibitors (HIVPIs) may have the potential to inhibit the Ddi1 in these parasites. Our previous experiments have proved HIVPIs can efficiently inhibit the Ddi1 activity of the tapeworm and have in vitro protoscolicidal activity. Here, we propose the hypothesis that HIVPIs are effective against echinococcosis. In this study, we plan to use the E. multilocularis larvae as a model to confirm the activity of HIVPIs versus the parasite and make the mechanism of treatment via inhibition of Ddi1 more clear, by using the methods, such as in vitro culture of E. multilocularis protoscoleces and infection in mice, and the techniques of drug screening, homologous gene knockout-complementation and transcriptome profiling. This study will provide novel insights for the development of new drugs against hydatid diseases.

棘球蚴病(包虫病)严重危害我国农牧区公共卫生安全，但迄今仅有苯并咪唑类药物可用于本病治疗。该类药物仅能抑制而不能有效杀死棘球蚴，急需开发新型治疗药物。申请者在棘球绦虫基因组上鉴定到具重要功能的DNA损伤诱导蛋白1（Ddi1）蛋白编码基因，发现此Ddi1与人类免疫缺陷病毒（HIV）蛋白酶三维结构竟然非常相似。推测HIV蛋白酶抑制剂（HIVPIs）可能会抑制棘球蚴Ddi1蛋白活性。我们前期实验已证明HIVPIs可有效抑制棘球蚴Ddi1功能，且在体外能有效杀伤棘球蚴原头节。由此提出艾滋病治疗药物HIVPIs有抗棘球蚴效果假说。本研究拟以多房棘球蚴为模型，通过原头节体外培养、小鼠体内感染等方法，利用药物筛选、同源基因敲除回补、转录组分析等技术，进一步明确HIVPIs的抗棘球蚴效果，并探究HIVPIs通过抑制Ddi1活性对棘球蚴产生杀伤作用的机理。本研究将为新型包虫病治疗药物的开发提供新思路。

棘球蚴病危害严重，但迄今仅有苯并咪唑类药物可用于本病治疗，急需开发新型治疗药物。申请者前期研究发现棘球绦虫的DNA损伤诱导蛋白1（Ddi1）蛋白，为潜在的HIV蛋白酶抑制剂（HIVPIs）靶标。本项目研究发现：经体外药物筛选，证明HIVPIs对多房棘球蚴的原头节的抑制效果，其中奈非那韦药效最好；经小鼠体内实验证明，奈非那韦在体内治疗棘球蚴包囊的效果明显，优于阿苯达唑；利用RNAi实验，证明Ddi1样蛋白为棘球蚴生存必须的关键基因，对棘球蚴原头节的蛋白外泌有显著抑制作用，敲低该基因后，寄生虫生存能力显著下降；经酶活性实验，首次证明绦虫Ddi1样蛋白具有明显的蛋白酶酶切活性，可以有效降解反转录型蛋白酶底物，其活性被可奈非那韦抑制，表明奈非那韦的作用机制与Ddi1样蛋白显著相关；转录组分析表明，奈非那韦对寄生虫产生的影响与Ddi1的作用机制类似；另外，基于全基因组及转录组等数据，鉴定到了多种潜在的抗寄生虫药物靶标。研究结果将为抗棘球蚴及其他寄生虫药物开发奠定基础。