Cysteine Proteases & Inhibitors in Nematode Development

描述（由申请人提供）：盘尾丝虫病，或河盲症，是 是一种主要的丝虫病，也是世界上第五大致盲原因。 世界控制程序的局限性和可能出现的 伊维菌素耐药菌株表明需要替代策略， 治疗和控制盘尾丝虫病。目前，很少有合适的目标， 化学疗法已被精确地确定在丝虫和其他寄生虫 线虫，部分原因是缺乏对基本生物学的了解， 这些寄生虫的生物化学。我们建议探索半胱氨酸蛋白酶， 化疗的潜在目标。在我们之前的研究中，我们发现了一种 半胱氨酸蛋白酶抑制剂、胰蛋白酶抑制剂和组织蛋白酶Z样半胱氨酸 蛋白酶，并提出，他们是必不可少的蜕皮，生长和 幼虫和蠕虫表皮的重塑，以及 微丝蚴最近，一种组织蛋白酶L样酶和另一种蛋白酶的成员， 半胱氨酸蛋白酶抑制剂家族被克隆，表明半胱氨酸蛋白酶和 它们的内源性抑制剂比最初认为的更复杂。然而，在这方面， 这些蛋白质在0.体内肠扭转，因为我们缺乏 用于观察寄生虫发育过程中基因表达的系统， 特别是在其成虫阶段和怀孕的雌蠕虫中。因为许多 线虫发育所必需的基因在自由生活和 寄生线虫，我们将利用同源的存在 C. elegans了解细胞的过程，其中0。 肠扭转半胱氨酸蛋白酶参与0. 人体内的肠扭转寄生虫综合利用遗传学， 分子、生物化学和解剖学研究，结合 定义明确的生物体，将有助于了解三种生物体的调节方式 不同的半胱氨酸蛋白酶对C. 优雅的0.肠扭转该项目有三个具体目标：1。 为了确定每种组织蛋白酶Z和 组织蛋白酶L样半胱氨酸蛋白酶在C. elegans的发展，然后 验证建议的0。肠扭转类似物将执行类似的功能 在0。肠扭转2.比较和对比了两种植物的生长发育规律， 组织特异性为0。volvulusand C.秀丽隐杆线虫胱抑素3.确定 底物特异性各为0。volvulusand C.线虫半胱氨酸蛋白酶和 在体外鉴定其特异性抑制剂。我们还将确定 分子量抑制剂，最终可以测试它们在 对成虫存活和微丝蚴发育的体内影响。这将 为开发有效药物、靶向半胱氨酸蛋白酶、 以控制盘尾丝虫病和丝虫病。

DESCRIPTION (provided by the applicant): Onchocerciasis, or river blindness, is a major filarial disease and is the fifth most common cause of blindness in the world. Limitations of control programs and the possible emergence of ivermectin-resistant strains suggest the need for alternative strategies for treatment and control of Onchocerciasis. Currently, few suitable targets for chemotherapy have been precisely identified in filarial and other parasitic nematodes, due in part to a lack of understanding of the basic biology and biochemistry of these parasites. We propose to explore cysteine proteases as potential targets for chemotherapy. In our previous studies we identified a cysteine protease inhibitor, onchocystatin, and a cathepsin Z-like cysteine protease, and proposed that they are essential for molting, growth and remodeling of the cuticle in larvae and adult worms, and the development of microfilariae. Recently, a cathepsin L-Iike enzyme and another member of the cystatin family were cloned, suggesting that the role of cysteine proteases and their endogenous inhibitors is more elaborate than initially thought. However, these proteins cannot be easily studied in 0. volvulus in vivo as we lack a system for observing gene expression during the development of the parasite, particularly in its adult stages and in the gravid female worms. As many of the essential genes for nematode development are conserved in free-living and parasitic nematodes, we will take advantage of the existence of homologous proteins in C. elegansto understand the cellular processes by which the 0. volvuluscysteine proteases participate in the development of the 0. volvulusparasite in humans. The integrated approach of using genetic, molecular, biochemical and anatomical studies in this proposal, combined with a well-defined organism, will result in understanding how regulation of three distinct cysteine proteases is critical for the development and survival of C. elegansand 0. volvulus. The proposed project has three specific objectives: 1. To establish the distinct physiological roles for each cathepsin Z and cathepsin L-like cysteine protease during C. elegans development and then verify that the proposed 0. volvulushomologues will perform similar functions in 0. volvulus. 2. To compare and contrast the developmental regulation and tissue specificity of 0. volvulusand C. elegans cystatins. 3. To determine the substrate specificity of each 0. volvulusand C. elegans cysteine protease and identify their specific inhibitors in vitro. We will also determine which low molecular weight inhibitors could, eventually, be tested for their in vivoeffects on adult worm survival and microfilariae development. This will provide the basis for developing effective drugs, targeting cysteine proteases, to control onchocerciasis and filariasis in the future.