Polysaccharide-based endotoxin antagonist for the treatment of sepsis

 描述（由申请人提供）：严重脓毒症和脓毒性休克是重症监护室经常遇到的危及生命的问题。目前基于抗生素治疗、正性肌力药、液体复苏和终末器官支持的治疗并不总是有效的，迫切需要新的治疗方法来降低脓毒症死亡率。一个正在进行的努力是去除或消除循环内毒素（或脂多糖，LPS），其主要负责在革兰氏阴性脓毒症中观察到的病理生理学紊乱。已经探索了可以通过抗原-抗体相互作用、静电相互作用或疏水相互作用与脂质A（充当LPS的毒性元素的阴离子糖脂）结合的几种化合物用于治疗脓毒症。然而，这些疗法几乎没有临床效果。困难部分来自于它们的疏水性和阳离子电荷，这是用于与LPS相互作用的特性，因为它们引起毒性和与蛋白质和其他血液成分的非特异性相互作用。 我们建议开发一种新的治疗剂，用于全身性脓毒症治疗的基础上，我们的壳聚糖衍生物，我们称之为两性离子壳聚糖（ZWC）。ZWC是通过低分子量壳聚糖（CS）的部分酰胺化产生的，但在电荷分布和水溶性方面与CS不同。与CS不同，ZWC带负电荷，在生理pH下是水溶性的，因此与血液成分相容。重要的是，它通过与LPS的细胞外相互作用抑制LPS攻击的巨噬细胞产生促炎细胞因子。此外，IP给药的ZWC减轻了LPS的全身作用或减弱了小鼠中LPS诱导的脓毒症的发作，而CS没有观察到不良组织反应的迹象。这些特征充分证明了ZWC作为新型LPS拮抗剂的研究。 我们的长期目标是在ZWC的基础上开发一种新的全身性脓毒症治疗方法。本研究的目的是产生在功效和效力方面与多粘菌素B相当或上级但无全身毒性的ZWC，并研究ZWC作为脓毒症的全身治疗的效用。我们的中心假设是，ZWC通过与LPS直接相互作用和与LPS/TLR 4信号传导途径的受体结合来抑制LPS活性;因此，ZWC修饰以增加这些特性将提高ZWC的效力和功效。为了检验这一假设，我们将用另外的羟基和疏水侧基修饰ZWC，并评估优化的ZWC的体外抗LPS活性，以其针对多粘菌素B为基准（目的1），以及优化的ZWC在严重脓毒症小鼠模型中作为独立疗法以及标准护理程序的补充的体内治疗效果（目的2）。预期所提出的工作将优化ZWC的性质以获得最佳治疗结果，并证明ZWC在全身治疗脓毒症中的有效性。本研究的成功完成将提供一种新的方法来中和LPS和降低脓毒症相关的死亡率，补充目前的脓毒症护理标准。

 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.