SERPIN STRUCTURE AND THE DEVELOPMENT OF NEUTROPHIL RESISTANT SERPINS

Serpin 结构和中性粒细胞抗性 Serpin 的开发

基本信息

批准号：
6397905
负责人：
SUSAN C BOCK
金额：
$ 16.54万
依托单位：
TEMPLE UNIV OF THE COMMONWEALTH
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-07-01 至 2001-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6397905
关键词：
antithrombin III chemical structure function disease /disorder model disseminated intravascular coagulation drug design /synthesis /production elastase inhibitor elastases endotoxins heparin laboratory rat protease inhibitor protein isoforms protein structure function septic shock serine proteinases

项目摘要

Granule, proteinases and oxidants relesd from activatedneutrophils during pathological inflammatory reactions contribute to the tissue destruction and circulatory collapse preceding organ failure in sepsis and ARDS. Vessel wall extracellular matrix and hemostatic pathway components are particularly sensitive to destruction by neutrophil elastase and cathepsin G (catG), and their degradation is associated with hypotension and disseminated intravascular coagulation (DIC). In Project 3 we will extend our serpin structure/funciton research twoards the practical goal of developing inhibitors which would be useful in sepsis and ARDS. Recent developments in antithrombin III (ATIII) research suggest potentially important funcitonal differences in the interactions of the naturally occurring alpha- and beta-ATIII isoforms with vessel wall heparan sulfate proteoglycans (HSPGs). This will be investigated in Aim 1 by determining isoform binding affinities for endothelial cells, and comparing their accessibility to fX-activating complex assembled on endothelial cells (which contain HSPGs) and on platelets and phospholipid vesicles (which do not). Aim 2 is to make vessel wall directed, elastase- and catG-resistant inhibitors of thrombin and fXa. Several animal and human studies have shown that high dose infusion of ATIII can reverse septic DIC and associated hypotension and organ failure. We hypothesize that high doses of ATIII are required due to (I) the preseence of an elastase cleavage site in the funcitonally important reactive loop of ATIII, and (ii) depletion of the high-heparin-affinity beta-ATIII isoform from commercial antithrombin concentrates. These considerations suggest that reduced amounts of a recombinant ATIII engineered to have enhanced heparin affinity and elastase- and catG- resistance may be effective for treating inflammatory DIC. Aim 3 is to make oxidation-resistant antielastase and anti-catG serpins that bind heparin. The heparin bidning property should increase association rates ofthesee inhibitors with elastase and cat G, and target them to the vessel wall where blocking extracellular matrix destruction and maintaining normal regulation of coagulation pathway assemblies is important. Finally, Airm 4 is to investigate the therapeutic potenital of the high-heparin-affinity neutrophil-resistant ATIII and the vessel wall directed anti-elastase and anti-catG serpins in an endotoxemic rat model of sepsis.

活化的中性粒细胞释放颗粒、蛋白酶和氧化剂在病理性炎症反应期间，有助于组织脓毒症器官衰竭前的破坏和循环衰竭和急性呼吸窘迫综合征。血管壁细胞外基质与止血途径成分对中性粒细胞的破坏特别敏感弹性蛋白酶和组织蛋白酶 G (catG)，它们的降解与伴有低血压和弥散性血管内凝血（DIC）。在项目3我们将扩展我们的丝氨酸蛋白酶抑制剂结构/功能研究两个标准开发抑制剂的实际目标将有助于败血症和急性呼吸窘迫综合征。抗凝血酶 III (ATIII) 的最新进展研究表明潜在的重要功能差异天然存在的 α- 和 β-ATIII 亚型的相互作用与血管壁硫酸乙酰肝素蛋白多糖（HSPG）。这将是在目标 1 中通过确定同种型结合亲和力进行研究内皮细胞，并比较它们对 fX 激活的可及性在内皮细胞（含有 HSPG）上组装的复合物血小板和磷脂囊泡（没有）。目标 2 是使血管壁定向、弹性蛋白酶和 catG 抗性凝血酶抑制剂和 fXa。多项动物和人类研究表明，高剂量输注 ATIII 可以逆转脓毒性 DIC 和相关的低血压和器官衰竭。我们假设高剂量的 ATIII 需要的原因是 (I) 中存在弹性蛋白酶切割位点 ATIII 功能上重要的反应环，以及 (ii) 的耗尽来自商业抗凝血酶的高肝素亲和力 β-ATIII 亚型集中。这些考虑因素表明，减少重组 ATIII 经改造具有增强的肝素亲和力弹性蛋白酶和 catG 抗性可能有效治疗炎症性 DIC。目标3是制造抗氧化抗弹性蛋白酶和结合肝素的抗catG丝氨酸蛋白酶抑制剂。肝素投标特性应增加这些抑制剂与弹性蛋白酶的关联率，并且 cat G，并将它们靶向阻断细胞外的血管壁基质破坏和维持凝血的正常调节通路组装很重要。最后，Airm 4 将调查高肝素亲和力中性粒细胞耐药性的治疗潜力 ATIII 和血管壁定向抗弹性蛋白酶和抗 catG 丝氨酸蛋白酶抑制剂脓毒症内毒素血症大鼠模型。