Novel Therapeutic Approach for Sepsis: Blocking the Vascular Response to Cytokine

描述(由申请人提供)：仅在美国，败血症每年就影响大约70万名患者，并导致超过21万人死亡。脓毒症是一种临床综合征，是宿主和感染性病原体复杂相互作用的结果，其特征是全身炎症和凝血级联反应的激活。血流动力学改变、广泛的微循环障碍和细胞改变导致血流和代谢需求之间的脱钩，参与了多器官功能障碍的发展，导致了大多数死亡。Navigen相信，它有一种独特的方法来防止与脓毒症相关的血液动力学变化和微循环障碍-血管泄漏。我们的科学联合创始人李迪安博士发现了一种新的受体Robo4，它在成熟的血管中表达，并在内皮损伤后上调。当被其蛋白质配体Slit2激活时，Robo4在体外和体内都能减少血管泄漏。我们的工作进一步表明，Robo4受体的激活干扰了包括肿瘤坏死因子-α、白介素2和凝血酶在内的多种通透性因子的下游信号级联反应。我们已经克隆了Slit2的活性片段Slit2N，它似乎与Slit2具有相同的药效，而且比天然蛋白更容易产生。Navigen已经证明，Slit2N在降低盲肠结扎和穿孔所致脓毒症动物模型中的死亡率方面是有效的。由于与蛋白质疗法相关的挑战，李博士试图识别一种可能以类似Slit2N的方式影响Robo4途径的小分子。李博士和他的合作者确定，Robo4激活的一个关键下游步骤是抑制一个小的GTP酶，ADP核糖化因子6(ARF6)。李博士和他的同事进一步确定，ARF6可以通过使用小分子抑制剂NAV838抑制其ARF-环境基金细胞内毒素2/ArNO而被阻断。李博士的实验室与Navigen合作，已经表明，通过阻断ARF-Global或通过Slit2N抑制Robo4激活ARF6，可以产生类似的表型：抑制血管内皮生长因子诱导的内皮细胞迁移。Navigen正在根据这项申请寻求资金，以确认Slit2N在治疗脓毒症方面的疗效，并确定NAV838作为一种小分子是否可能是一种比Slit2N更可行的治疗败血症的方法。纳维根预计将把这种高级化合物向前推进到IND，最终实现商业化。本研究旨在实现三个目标：1)建立肺炎假单胞菌肺炎(PAP)引起的脓毒症动物模型，在该动物模型中，抗生素治疗剂量和时间被滴定，以达到约50%的7天死亡率；2)在CLP和PAP模型中测试Slit2N的疗效并确定最有效的联合治疗剂量(PAP在Aim 1中建立)；3)通过在CLP和PAP动物模型上的试验，确定NAV838联合抗生素是否可能比Slit2N+抗生素有更好的疗效。 公共卫生相关性：脓毒症是一种临床综合征，是宿主和感染性病原体之间复杂相互作用的结果，其特征是全身激活多种炎症途径，包括细胞因子网络和凝血。与严重脓毒症相关的死亡率在30%到50%之间，而脓毒症是外科患者和创伤受害者发病率和死亡率的主要原因。目前，还没有有效的治疗方法来治疗脓毒症。

DESCRIPTION (provided by applicant): In the United States alone, sepsis affects approximately 700,000 patients and leads to death of over 210,000 people per year. Sepsis is a clinical syndrome that results from a complex interaction between host and infectious agents, and is characterized by systemic activation of inflammatory and coagulation cascades. Hemodynamic changes, widespread microcirculatory disturbances and cellular alterations, leading to an uncoupling between blood flow and metabolic requirements, are implicated in the development of multiple organ dysfunction, responsible for most of deaths. Navigen believes that it has a unique means of preventing the hemodynamic changes and microcirculatory disturbances - the vascular leak - associated with sepsis. Our scientific co-founder, Dr. Dean Li, identified a novel receptor, Robo4, that is expressed in mature vessels and is upregulated following endothelial injury. Robo4, when activated by its protein ligand, Slit2, reduces vascular leak in vitro and in vivo. Our work has further illustrated that activation of the Robo4 receptor interferes with the downstream signaling cascades from multiple permeability factors including TNF-alpha, interleukins, and thrombin. We have cloned an active fragment of Slit2, Slit2N, which appears to have identical efficacy to Slit2, and is more easily produced than the native protein. Navigen has demonstrated that Slit2N is effective in reducing mortality in the animal model of sepsis induced by cecal ligation and puncture. Due to the challenges associated with protein therapeutics, Dr. Li sought to identify a small molecule that could affect the Robo4 pathway in a manner similar to Slit2N. Dr. Li and collaborators determined that a critical downstream step in Robo4 activation is the inhibition of a small GTPase, ADP ribosylation factor 6 (ARF6). Dr. Li and colleagues further determined that ARF6 can be blocked through inhibition of its ARF-GEF, cytohesin2/ARNO, using a small molecule inhibitor, NAV838. Dr. Li's laboratory, in collaboration with Navigen, has shown that inhibition of ARF6, either by blocking its ARF-GEF or through the inhibitory activation of Robo4 by Slit2N, produced similar phenotypes: inhibition of VEGF-induced migration in endothelial cells. Navigen is seeking funding under this application to confirm the efficacy of Slit2N in the treatment of sepsis and to determine whether NAV838, as a small molecule, may be an even more viable approach to treating sepsis than Slit2N. Navigen would anticipate advancing the superior compound forward to IND for eventual commercialization. The proposed research is intended to accomplish 3 goals: 1) develop animal models of sepsis induced by pseudomonas pneumonia (PAP) in which antibiotic therapy dose and timing are titrated to achieve seven-day mortality of approximately 50%, 2) test efficacy and indentify most efficacious dose of Slit2N combined with antibiotic therapy in models of CLP and PAP (PAP established in Aim 1), 3) determine whether NAV838 combined with antibiotics may have greater efficacy than Slit2N plus antibiotics by testing in animal models of CLP and PAP. PUBLIC HEALTH RELEVANCE: Sepsis is a clinical syndrome that results from a complex interaction between host and infectious agents, and is characterized by systemic activation of multiple inflammatory pathways, including cytokine network and coagulation. Mortality associated with severe sepsis ranges between 30 to 50 percent and sepsis is the leading cause of morbidity and mortality in surgical patients and trauma victims. Currently, there are no effective therapies for the treatment of Sepsis.