Polysaccharide-based endotoxin antagonist for the treatment of sepsis

 描述(由申请人提供)：严重败血症和感染性休克是重症监护病房经常遇到的危及生命的问题。目前基于抗生素治疗、肌松、液体复苏和终末器官支持的治疗方法并不总是有效的，迫切需要新的治疗方法来降低脓毒症的死亡率。一项正在进行的努力是清除或灭活循环内毒素(或脂多糖)，它是革兰氏阴性脓毒症中出现的病理生理紊乱的主要原因。几种化合物可以通过抗原-抗体相互作用、静电相互作用或疏水相互作用与脂类A(一种阴离子糖脂)结合，用于治疗脓毒症。然而，这些疗法在临床上收效甚微。困难的部分原因是它们的疏水性和阳离子电荷，这正是与内毒素相互作用所用的性质，因为它们导致与蛋白质和其他血液成分的毒性和非特异性相互作用。我们建议在我们的壳聚糖衍生物的基础上开发一种新的治疗全身脓毒症的药物，我们称之为两性离子壳聚糖(ZWC)。ZWC是由低分子量壳聚糖(CS)部分酰胺化而成，但其电荷分布和水溶性与CS不同。与CS不同的是，ZWC带负电荷，在生理pH下溶于水，因此与血液成分兼容。重要的是，它通过细胞外与内毒素的相互作用，抑制内毒素攻击的巨噬细胞产生促炎细胞因子。此外，腹腔注射ZWC减轻了内毒素的全身效应或减轻了内毒素诱导的小鼠脓毒症的发病，CS没有看到不良组织反应的迹象。这些特点充分证明了ZWC作为一种新型的内毒素拮抗剂的研究价值。我们的长期目标是在ZWC的基础上开发一种新的全身治疗脓毒症的方法。本研究的目的是制备一种在疗效和效力方面与多粘菌素B相当或优于多粘菌素B但无全身毒性的ZWC，并探讨ZWC作为全身治疗脓毒症的用途。我们的中心假设是ZWC通过与内毒素的直接相互作用和与LPS/TLR4信号通路的受体结合来抑制内毒素的活性；因此，对ZWC进行修饰以增加这些特性将提高ZWC的效力和疗效。为了验证这一假说，我们将用额外的羟基和疏水悬浮物来修饰ZWC，并评估其以多粘菌素B为基准的优化ZWC的体外抗内毒素活性(目标1)，以及作为标准护理程序的补充及单独治疗的优化ZWC在严重脓毒症小鼠模型中的体内治疗效果(目标2)。这项拟议的工作有望优化ZWC的性能，以获得最佳治疗结果，并证明ZWC在全身治疗脓毒症方面的有效性。这项研究的成功完成将为中和内毒素和降低脓毒症相关死亡率提供一种新的方法，补充目前脓毒症的护理标准。

 DESCRIPTION (provided by applicant): Severe sepsis and septic shock are life-threatening problems frequently encountered in the critical care unit. Current treatments based on antibiotic therapy, inotropes, fluid resuscitation, and end-organ support are not always effective, and there is an urgent need for new therapies to reduce sepsis mortality. One ongoing effort is to remove or inactivate circulating endotoxins (or lipopolysaccharides, LPS), which are primarily responsible for the pathophysiological derangements seen in gram-negative sepsis. Several compounds that can bind to lipid A, an anionic glycolipid serving as a toxic element of LPS, via antigen-antibody interactions, electrostatic interactions, or hydrophobic interactions have been explored for the treatment of sepsis. However, these therapies have made little clinical impacts. The difficulties partly arise from their hydrophobicity and cationic charge, the very properties used for interaction with LPS, as they cause toxicity and non-specific interactions with proteins and other blood components. We propose to develop a new therapeutic agent for systemic sepsis treatment based on our chitosan derivative, which we call zwitterionic chitosan (ZWC). ZWC is created by partial amidation of low molecular weight chitosan (CS) but distinct from CS by the charge profile and water solubility. Unlike CS, ZWC is negatively charged and water-soluble at physiological pH and thus compatible with blood components. Importantly, it suppresses the production of pro-inflammatory cytokines by LPS-challenged macrophages via extracellular interaction with LPS. Moreover, IP-administered ZWC mitigated systemic effect of LPS or attenuated the onset of LPS-induced sepsis in mice, with no signs of adverse tissue responses seen with CS. These features well justify the investigation of ZWC as a novel LPS antagonist. Our long-term goal is to develop a new systemic treatment for sepsis based on ZWC. The objective of this study is to produce a ZWC, comparable or superior to polymyxin B in efficacy and potency but with no systemic toxicities, and to investigate the utility of ZWC as a systemic therapy of sepsis. Our central hypothesis is that ZWC suppresses LPS activity via direct interaction with LPS and binding to receptors of LPS/TLR4 signaling pathways; therefore, ZWC modifications to increase these properties will improve the potency and efficacy of ZWC. To test this hypothesis, we will modify ZWC with additional hydroxyl groups and hydrophobic pendants and evaluate its in vitro anti-LPS activity of optimized ZWC benchmarking it against polymyxin B (Aim 1) and in vivo therapeutic effects of the optimized ZWC in a mouse model of severe sepsis as a standalone therapy as well as a supplement to standard care procedure (Aim 2). The proposed work is expected to optimize the properties of ZWC for best therapeutic outcomes and prove the effectiveness of ZWC in systemic treatments of sepsis. Successful accomplishment of this study will provide a new way to neutralize LPS and reduce the sepsis-related mortality, complementing the current standard of care of sepsis.