Mitochondrial-Targeted Therapeutics for Treatment of Spinal Cord Injury

描述（由申请人提供）：脊髓损伤（SCI）在美国是一个长期的医疗保健问题，除了适度有效的甲基强的松龙外，目前临床上没有可用于治疗急性SCI的神经保护干预措施。我们发表的数据和初步结果表明，关键线粒体酶的氧化损伤和随后的线粒体功能障碍是脊髓损伤后神经病理后遗症的关键。这一建议的重点是直接针对线粒体功能障碍作为一种新的治疗干预手段，其基本概念是SCI诱导的兴奋性毒性增加线粒体Ca2+循环/过载和活性氧（ROS）的产生，最终导致线粒体功能障碍和谷胱甘肽（GSH）消耗。我们的方法是双管齐下的，旨在利用一种新的细胞渗透抗氧化剂和谷胱甘肽前体，NACA （n -乙酰半胱氨酸的酰胺形式），以及一种用于能源生产的替代生物燃料底物，乙酰左肉碱（ALC），在SCI后减少线粒体ROS的产生。我们已发表的和初步的数据表明，NACA和ALC都能改善大鼠挫伤性脊髓损伤后的线粒体生物能量，并且延长NACA或ALC治疗可增加损伤后的组织保留。计划中的实验旨在验证一种新的假设，即减少对关键线粒体蛋白的氧化损伤将维持线粒体生物能量，从而增加脊髓挫伤后的神经保护和改善功能恢复。具体来说，我们将：1)表征参与生物能量学的特定线粒体蛋白的氧化损伤，并验证NACA治疗改善脊髓损伤后线粒体氧化损伤的假设。2)验证NACA和ALC联合治疗将协同作用于脊髓损伤后线粒体稳态的假设。3)验证NACA和ALC联合治疗将增加脊髓损伤后组织保留和促进长期功能恢复的假设。至关重要的是，该应用是围绕我们开发的几种新技术建立的，这些技术用于从受损脊髓中分离突触（神经元）和非突触（体细胞和胶质）线粒体，以及L1/L2挫伤SCI范例，该范例证明了神经保护与后肢功能恢复的显着改善之间的显着相关性。总的来说，拟议的实验将确定关键的线粒体事件，这些事件可能成为药物干预的潜在新靶点，以更有效地治疗脊髓损伤，也许还有其他中枢神经系统损伤。

DESCRIPTION (provided by applicant): Spinal cord injury (SCI) is a long-term health care problem in the United States, and with the exception of the modestly effective methylprednisolone, there is currently no neuroprotective intervention clinically available for treatment of acute SCI. Our published data and preliminary results demonstrate that oxidative damage to key mitochondrial enzymes and subsequent mitochondrial dysfunction is key to the neuropathological sequalae following SCI. This proposal focuses on directly targeting mitochondrial dysfunction as a novel therapeutic intervention for contusion SCI, the fundamental concept being that SCI-induced excitotoxicity increases mitochondrial Ca2+ cycling/overload and the production of reactive oxygen species (ROS), ultimately leading to mitochondrial dysfunction and glutathione (GSH) depletion. Our approach is two-pronged, aimed at reducing mitochondrial ROS production utilizing a novel, cell-permeant antioxidant and GSH precursor, NACA (the amide form of N-acetylcysteine), as well as an alternative biofuel substrate for energy production, acetyl-l-carnitine (ALC), following SCI. Our published and preliminary data signify that both NACA and ALC improve mitochondrial bioenergetics following contusion SCI in rats, and that prolonged NACA or ALC treatment increases tissue sparing following injury. The planned experiments are designed to test the novel hypothesis that reducing oxidative damage to key mitochondrial proteins will maintain mitochondrial bioenergetics, thus leading to increased neuroprotection and improved functional recovery following contusion SCI. Specifically we will: 1) Characterize oxidative damage to specific mitochondrial proteins involved in bioenergetics and test the hypothesis that NACA treatment ameliorates mitochondrial oxidative damage following SCI, 2) Test the hypothesis that a combinatorial treatment with NACA and ALC will act synergistically to preserve mitochondrial homeostasis following SCI, and 3) Test the hypothesis that a combinatorial treatment with NACA and ALC will increase tissue sparing and promote long-term functional recovery following SCI. Critically, this application is built around the utilization of several novel techniques we have developed for isolating synaptic (neuronal) and non-synaptic (soma and glia) mitochondria from the injured spinal cord, as well as an L1/L2 contusion SCI paradigm that demonstrates a significant correlation between neuroprotection and remarkable improvements in recovery of hind limb function. Collectively, the proposed experiments will pinpoint key mitochondrial events that could be potential novel targets for pharmacological interventions to more effectively treat SCI and, perhaps, other CNS injuries.