TETANUS TOXIN C-FRAGMENT/SOD HYBRID AS A NEUROPROTECTANT

破伤风毒素 C 片段/SOD 混合物作为神经保护剂

• 批准号: 6098635
• 负责人: PAUL S FISHMAN
• 金额: $ 17.77万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2000-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6098635
• 关键词: active immunization; antibacterial antibody; antioxidants; central nervous system; chimeric proteins; cytotoxicity; drug administration routes; drug vehicle; embryo /fetus tissue /cell culture; enzyme activity; free radicals; glutamates; human tissue; laboratory mouse; laboratory rat; neuropharmacology; neuroprotectants; neurotoxins; oxidative stress; pharmacokinetics; recombinant proteins; superoxide dismutase; tetanus toxin; tetanus toxoid; tissue /cell culture

The enzyme superoxide dismutase (SOD) is under active investigation as a form of neuroprotective therapy in both neurodegenerative diseases and brain injury. Increasing cellular levels of SOD have increased neuronal viability in several models of free radical mediated injury. Poor penetration of SOD into the nervous system limits its utility as a potential therapeutic. The non-toxic binding fragment of tetanus toxin, termed C-fragment (CF), shares many characteristics of its parent protein, such as entry into and persistence within the nervous system. These properties have led investigators to demonstrate its ability to be used as a vector to deliver large proteins including enzymes to the nervous system in experimental animals. Using recombinant genetic methods we have produced a hybrid protein containing CF and SOD. This protein retains the neuronal binding properties of CF and the enzymatic properties of SOD. This proposal will explore the potential of this hybrid protein as a novel neuroprotectant. Using both a cell line and cultured embryonic neurons we will determine to what extent CF-SOD can raise cellular levels of SOD. We will then determine if cells treated with CF-SOD are resistant to damage and death due to exposure to toxic levels of the excitatory amino acid glutamate. In animals we will determine the extent to which CF-SOD enhances the nervous system uptake SOD. Using radiolabeled proteins we will compare uptake of native SOD and CF-SOD after intravenous, intramuscular and intrathecal administration. We will determine the amount of distribution and lifetime of these radiolabeled proteins within the nervous system. The effect of anti-tetanus antibodies on nervous system uptake of CF-SOD will be examined in both previously immunized rats and rats receiving sera from immunized humans. SOD levels will be measured enzymatically within the nervous system after administration of CF-SOD. We will determine to what extent CF-SOD can raise cellular levels of SOD. Using subcellular fractionation techniques, we will determine what cellular compartment accumulates SOD activity of administration of CF-SOD. These experiments will explore the biology of this unique protein in the nervous system and evaluate its potential as a neuro-protectant. Successful completion of this proposal will provide the basis for preclinical nd clinical study of CF-SOD as a new therapeutic agent for neurological disease.

超氧化物歧化酶（SOD）正在积极研究中， 在神经退行性疾病和 脑损伤 细胞内SOD水平的增加， 在几种自由基介导的损伤模型中的活力。 贫困 SOD对神经系统的渗透限制了其作为抗氧化剂的效用。 潜在的治疗 破伤风毒素的无毒结合片段， 称为C片段（CF），与其亲本具有许多特征 蛋白质，如进入和持久性内的神经系统。 这些特性使研究人员证明了它的能力， 用作载体，将包括酶在内的大蛋白质递送到 实验动物的神经系统 利用重组基因 方法我们已经产生了含有CF和SOD的杂合蛋白。 这 蛋白质保留了CF和酶的神经元结合特性， SOD的特性 该提案将探讨这一潜力， 杂合蛋白作为新型神经保护剂。 使用细胞系和 培养胚胎神经元，我们将确定CF-SOD在多大程度上可以 提高细胞SOD水平。 然后我们将确定细胞处理 具有CF-SOD的细胞可以抵抗由于暴露于有毒物质而引起的损伤和死亡。 兴奋性氨基酸谷氨酸的水平。 在动物中，我们将 确定CF-SOD增强神经系统摄取的程度 该死 使用放射性标记的蛋白质，我们将比较天然SOD的摄取 静脉注射、肌肉注射和鞘内注射后， 局 我们将确定分配的数量， 这些放射性标记的蛋白质在神经系统中的寿命。 的 抗破伤风抗体对神经系统摄取CF-SOD影响 在先前免疫的大鼠和接受血清的大鼠中进行检查 免疫的人类。 SOD水平将在 CF-SOD给药后的神经系统。 我们将决定 CF-SOD能在多大程度上提高细胞内SOD水平。 使用亚细胞 分离技术，我们将确定哪些细胞区室 累积施用CF-SOD的SOD活性。 这些实验 将探索神经系统中这种独特蛋白质的生物学， 评估其作为神经保护剂的潜力。 成功完成 为临床前和临床研究提供了依据 CF-SOD作为一种新的神经系统疾病的治疗剂。