Pn3Pase as an Effective Enzyme Therapeutic for Streptococcus Pneumoniae type 3 Infection

项目摘要 肺炎链球菌(Spn)仍然是最致命的感染性病原体之一，导致超过一年的 全球每年有100万人死亡，占#年继发性感染的大部分 免疫功能受损的病人。与这种病原体相关的主要毒力机制之一是它的 多糖胶囊，决定细菌的成功定植和感染，以及 作为一种防御机制来抵抗免疫介导的病原体杀灭。SPN血清3型(Spn3)是 被认为是Spn毒力最强的血清型之一，尽管目前的多价疫苗包括 结合抗Spn3的靶点，这种血清型在Spn感染病例中的总体比例实际上 在过去的十年里有所增加。此外，Spn3与抗生素耐药性的增加直接相关。 因此，Spn3影响的严重性，以及设计针对它的治疗方法所产生的困难，不能 言过其实。肺炎酶(Pn3Pase)是一种酶产物，我们以前已经证明它是高度 能有效降解3型Spn的荚膜多糖。我们的体外研究表明，治疗 带有肺炎酶的Spn3会使细菌更容易受到吞噬细胞的杀伤机制的影响。 此外，受到Spn3致命挑战的小鼠可以在使用肺炎酶治疗后获救： 接受肺炎酶治疗的小鼠100%存活，而接受肺炎酶治疗的小鼠100% 无活性的酶或载体在Spn3致命攻击后的两天内死亡。这些结果表明，肺炎酶 在使Spn3对宿主免疫机制敏感方面具有极高的效率，该酶具有显著的 有潜在的治疗作用。作为Patiotactix，LLC的一部分，我们对该项目的目标是： 目的1：研究该酶的药代动力学、潜在的免疫原性和细胞毒性。 目的2：阐明该酶对Spn3感染的治疗价值。 在我们发表的研究中，我们使用了天然的全长蛋白质产品来降解One的胶囊 Spn3强毒株效果显著，并进一步表明该酶产物不诱导细胞 体外和体内实验中对宿主哺乳动物细胞的毒性。我们在这方面的近期目标 因此，项目是扩大这些研究，以测试药代动力学，评估酶对 更多的血清3型Spn菌株，并进行持续的实验以证实其治疗价值 肺炎酵母菌和同时缺乏对给药的非靶向影响。的长期目标 肺炎酶将通过在高等动物系统中测试其有效性来确立其作为治疗剂(即， 拥有大量Spn 3型临床分离株的灵长类动物)，并进入临床试验 开发这种酶作为一种高度相关的治疗方法来对抗这种极其致命的病原体。

Project Summary Streptococcus pneumoniae (Spn) remains one of the deadliest infectious agents responsible for more than a million deaths worldwide per year as well as making up a majority of secondary infections in immunocompromised patients. One of the major virulence mechanisms associated with this pathogen is its polysaccharide capsule, which determines successful colonization and infection of the bacterium as well as serving as a defense mechanism against immune-mediated killing of the pathogen. Spn serotype 3 (Spn3) is considered one of the most virulent serotypes of Spn, and despite current multivalent vaccines that include conjugate targets against Spn3, the overall proportion of this serotype in Spn infection cases has actually increased over the last ten years. Furthermore, Spn3 is directly associated with increased antibiotic resistance. Thus, the severity of the impact of Spn3, as well as the difficulties arising in designing therapies against it, cannot be overstated. Pneumozyme (Pn3Pase) is an enzyme product that we have previously shown to be highly effective in degrading the capsular polysaccharide of type 3 Spn. Our in vitro studies have shown that treating Spn3 with Pneumozyme will render the bacterium more susceptible to killing mechanisms by phagocytic cells. Furthermore, mice that are lethally challenged with Spn3 can be rescued following treatment with Pneumozyme: 100% of mice receiving Pneumozyme treatment survival the challenge whereas 100% of mice that receive inactive enzyme or vehicle die within two days of Spn3 lethal challenge. These results indicate that Pneumozyme has extremely high efficacy in sensitizing Spn3 to host immune mechanisms and that the enzyme has remarkable potential as a therapeutic. As part of Pneumotactix, LLC, our aims for this project are thus: Aim 1: Characterize the pharmacokinetics, potential immunogenicity and cytotoxicity of the enzyme. Aim 2: Elucidate the therapeutic value of the enzyme against Spn3 infection. In our published studies, we have used the native, full-length protein product to degrade the capsule of one virulent strain of Spn3 to great effect, and have further shown that this enzyme product does not induce cellular toxicity against host mammalian cells in both in vitro and in vivo experiments. Our immediate goals for this project are therefore to expand these studies to test pharmacokinetics, assess enzyme efficacy against additional strains of serotype 3 Spn, and perform continued experiments to confirm the therapeutic value of Pneumozyme and concurrent lack of off-target effects upon administration. The long-term goals for Pneumozyme are to establish it as a therapeutic agent by testing its efficacy in higher animal systems (i.e., primates) with significant sample numbers of Spn type 3 clinical isolates, and move towards clinical trials to develop this enzyme as a highly relevant therapeutic against this incredibly virulent pathogen.