Combination drug treatment to inhibit multiple cell death pathways after TBI

抑制 TBI 后多种细胞死亡途径的联合药物治疗

• 批准号: 7985713
• 负责人: ALAN Ira FADEN
• 金额: $ 48.62万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-11-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7985713
• 关键词: Cell Death; Drug Combinations; Pathway interactions

DESCRIPTION (provided by applicant): This application entitled "Attenuation of Diverse Cell Death Pathways after Traumatic Brain Injury by Multi- drug Combination Therapy" addresses Broad Challenge Area (15): Translational Science and Specific Challenge Topic: 15-HD-104, Multi-drug Combination Therapy for TBI and Stroke Treatment. The purpose of the proposed studies is to evaluate a novel combination treatment strategy in experimental traumatic brain injury (TBI). Traditional neuroprotective treatment strategies for TBI aim to prevent delayed (secondary) neuronal cell death, generally by inhibiting one proposed cell death mechanism. Yet considerable research indicates that multiple pathways and mechanisms of cell death contribute to tissue loss. In focal TBI, the central injury site is thought to largely reflect necrotic cell death, which primarily occurs within the first 6-8 hours after trauma and is associated with severe bioenergetic compromise. Our recent work, however, indicates that in several rodent TBI models -mouse controlled cortical impact (CCI) or rat lateral fluid percussion (LFP)- cell death in the central core region also includes a substantial component of caspase-independent apoptosis, whereas the better known caspase-dependent cell death is detected in the more peripheral regions where ATP and ADP levels are largely preserved. The caspase-independent programmed cell death (PCD) is due in large part to the activation/translocation of apoptosis-inducing factor (AIF). Such cell death occurs relatively late after injury (24-72h) and can be inhibited by delayed treatment hours after the insult. These data raise serious questions about classical assumptions regarding mechanism of post-traumatic brain injury, suggesting the possibility of newer treatment approaches with extended therapeutic windows and explaining why, if used exclusively, caspase inhibitors may have only a partial protective effect. Ideally, treatment for TBI should attempt to inhibit both caspase-independent and caspase-dependent PCD. Recent studies in ischemia, as well as unpublished work from our laboratory, suggest two intriguing approaches for limiting the three major pathways of cell death after injury. One approach is to up-regulate heat shock protein 70 (HSP70), which binds both Apaf-1 and AIF at distinct sites, thereby neutralizing their pro- apoptotic functions by preventing the formation of the apoptosome (and caspase 3) and attenuating AIF mediated actions. Our preliminary data show that TBI causes up-regulation of HSP70 in many neurons within the injury zone; those neurons expressing HSP 70 show neither caspase-3 activation nor AIF translocation. Up-regulation of HSP70 in cerebral ischemia is strongly protective. The other approach is to inhibit PARP-1. PARP-1, through the release of poly ADP ribose (PAR), is a critical upstream activator AIF release from neuronal mitochondria; it also more recently has been shown to be a critical activator of microglia. Inhibition of PARP-1 after TBI strongly attenuates both caspase dependent and independent forms of PCD, as well as microglial activation, with markedly improved outcome. The advantage of each of these strategies is that their therapeutic window should be very broad, at least 24h. By combining these distinct therapeutic strategies, additive or synergistic protective effects may potentially be achieved. Should our hypotheses be supported, concepts regarding treatment of TBI will be markedly altered and target populations for therapeutic intervention considerably expanded. Combination treatment evaluation is proposed in two pathobiologically different models in different species, with the assumption that potent treatment effects duplicated across models and species makes ultimate clinical translation more likely. Specific hypotheses include: 1) HSP-70 inducers or PARP-1 inhibitors attenuate caspase-independent and caspase-dependent PCD after TBI, reducing long-term neurological dysfunction; 2) Each treatment approach has a long therapeutic window of at least 24h; 3) combined therapy with HSP-70 inducers and PARP-1 inhibitors demonstrate additive and/or synergistic effects in both mouse CCI and rat LFP models. We propose the following specific aims: 1) to compare the efficacy, dose response, and therapeutic window of two structurally distinct HSP-70 inducers with regard to attenuation of post-traumatic neuronal cell death and improved functional recovery after moderate CCI injury in mice; 2) to compare the efficacy, dose response and therapeutic window of two structurally distinct PARP-1 inhibitors with regard to attenuation of post-traumatic neuronal cell death and improved functional recovery after moderate CCI injury in mice; 3) to determine whether combined multi-drug therapy with the best HSP-70 inducer and best PARP-1 inhibitor, at optimal doses, has additive or synergistic effects on cell death, microglial activation, and neurodegenerative conditions up to 3 months post-injury. The Centers for Disease Control and Prevention (CDC) defines traumatic brain injury (TBI) as craniocerebral trauma associated with a decreased level of consciousness, amnesia, other neurologic or neuropsychological abnormalities, skull fracture, intracranial lesions, or death. It has been reported that the combined incidence of fatal and hospitalized TBI among all age groups has a median annual incidence of 101 per 100,000. Approximately 20% of TBIs cause death either immediately or during acute hospital care, with estimated annual rates of mild TBI treated only in outpatient facilities or hospital emergency departments (EDs) in the United States were 392 and 540 visits per 100,000, respectively. A disability prevalence of 37% has been reported for TBI patients followed more than one year after hospitalization; based on this figure, the CDC has estimated that nearly 2% of the entire US population has TBI-related disabilities. Traumatic brain injury (TBI) may occur in as many as 22% of troops deployed in Afghanistan and Iraq. Recent studies in ischemia, as well as unpublished work from our laboratory, suggest two intriguing approaches for limiting the three major pathways of cell death after injury. One approach is to up-regulate heat shock protein 70 (HSP70), which binds both Apaf-1 and AIF at distinct sites, thereby neutralizing their pro-apoptotic functions by preventing the formation of the apoptosome (and caspase 3) and attenuating AIF mediated actions. Our preliminary data show that TBI causes up- regulation of HSP70 in many neurons within the injury zone; those neurons expressing HSP 70 show neither caspase-3 activation nor AIF translocation. Up-regulation of HSP70 in cerebral ischemia is strongly protective. The other approach is to inhibit PARP-1. PARP-1, through the release of poly ADP ribose (PAR), is a critical upstream activator AIF release from neuronal mitochondria; it also more recently has been shown to be a critical activator of microglia. Inhibition of PARP-1 after TBI strongly attenuates both caspase dependent and independent forms of PCD, as well as microglial activation, with markedly improved outcome. The advantage of each of these strategies is that their therapeutic window should be very broad, at least 24h. By combining these distinct therapeutic strategies, additive or synergistic protective effects may potentially be achieved. Should our hypotheses be supported, concepts regarding treatment of TBI will be markedly altered and target populations for therapeutic intervention considerably expanded. Combination treatment evaluation is proposed in two pathobiologically different models in different species, with the assumption that potent treatment effects duplicated across models and species makes ultimate clinical translation more likely.

描述（由申请人提供）： 本申请题为“通过多药物组合疗法减弱创伤性脑损伤后多种细胞死亡途径”，解决了广泛的挑战领域（15）：转化科学和具体挑战主题：15-HD-104，TBI 和 TBI 的多药物组合疗法中风治疗。拟议研究的目的是评估实验性脑外伤（TBI）的新型联合治疗策略。 TBI 的传统神经保护治疗策略旨在防止延迟性（继发性）神经元细胞死亡，通常是通过抑制一种拟议的细胞死亡机制。然而大量研究表明，细胞死亡的多种途径和机制都会导致组织损失。在局灶性 TBI 中，中央损伤部位被认为在很大程度上反映了坏死性细胞死亡，这主要发生在创伤后的前 6-8 小时内，并与严重的生物能损害相关。然而，我们最近的工作表明，在几种啮齿动物 TBI 模型中 - 小鼠控制的皮质冲击 (CCI) 或大鼠侧向液体冲击 (LFP) - 中央核心区域的细胞死亡也包括不依赖 caspase 的细胞凋亡的重要组成部分，而更广为人知的 caspase 依赖性细胞死亡是在 ATP 和 ADP 水平基本保持不变的更外围区域中检测到的。不依赖半胱天冬酶的程序性细胞死亡（PCD）在很大程度上是由于凋亡诱导因子（AIF）的激活/易位。这种细胞死亡在损伤后发生得相对较晚（24-72小时），并且可以通过损伤后延迟治疗数小时来抑制。这些数据对关于创伤后脑损伤机制的经典假设提出了严重的质疑，表明了具有延长治疗窗的新治疗方法的可能性，并解释了为什么如果单独使用 caspase 抑制剂可能只具有部分保护作用。理想情况下，TBI 的治疗应尝试抑制 caspase 依赖性和 caspase 依赖性 PCD。 最近对缺血的研究以及我们实验室未发表的工作提出了两种有趣的方法来限制损伤后细胞死亡的三种主要途径。一种方法是上调热休克蛋白 70 (HSP70)，该蛋白在不同位点结合 Apaf-1 和 AIF，从而通过阻止凋亡体（和 caspase 3）的形成并减弱 AIF 介导的作用来中和它们的促凋亡功能。行动。我们的初步数据表明，TBI 会导致损伤区内许多神经元的 HSP70 上调；那些表达 HSP 70 的神经元既没有表现出 caspase-3 激活，也没有表现出 AIF 易位。 HSP70 在脑缺血中的上调具有很强的保护作用。另一种方法是抑制 PARP-1。 PARP-1 通过释放聚 ADP 核糖 (PAR)，是神经元线粒体释放 AIF 的关键上游激活剂；最近它还被证明是小胶质细胞的关键激活剂。 TBI 后抑制 PARP-1 可强烈减弱 caspase 依赖性和非依赖性 PCD 形式以及小胶质细胞的激活，从而显着改善预后。这些策略的优点是它们的治疗窗应该非常宽，至少 24 小时。通过结合这些不同的治疗策略，可能会实现相加或协同的保护作用。如果我们的假设得到支持，关于 TBI 治疗的概念将显着改变，治疗干预的目标人群将大大扩大。在不同物种的两个病理生物学不同模型中提出了联合治疗评估，假设跨模型和物种复制有效的治疗效果使得最终的临床转化更有可能。 具体假设包括：1) HSP-70 诱导剂或 PARP-1 抑制剂可减弱 TBI 后不依赖 caspase 和依赖 caspase 的 PCD，从而减少长期神经功能障碍； 2）每种治疗方法都有至少24小时的长治疗窗； 3) HSP-70诱导剂和PARP-1抑制剂的联合治疗在小鼠CCI和大鼠LFP模型中显示出相加和/或协同效应。 我们提出以下具体目标：1) 比较两种结构不同的 HSP-70 诱导剂在减轻小鼠中度 CCI 损伤后创伤后神经元细胞死亡和改善功能恢复方面的功效、剂量反应和治疗窗； 2) 比较两种结构不同的 PARP-1 抑制剂在减轻小鼠中度 CCI 损伤后创伤后神经元细胞死亡和改善功能恢复方面的功效、剂量反应和治疗窗； 3) 确定最佳剂量的最佳 HSP-70 诱导剂和最佳 PARP-1 抑制剂的多药联合治疗是否对损伤后 3 个月内的细胞死亡、小胶质细胞激活和神经退行性疾病具有累加或协同作用。美国疾病控制与预防中心 (CDC) 将创伤性脑损伤 (TBI) 定义为与意识水平下降、健忘症、其他神经或神经心理异常、颅骨骨折、颅内病变或死亡相关的颅脑损伤。据报道，所有年龄组中致命性和住院性 TBI 的年发病率中位数为每 100,000 人 101 例。大约 20% 的 TBI 导致立即死亡或在急性住院治疗期间死亡，估计仅在美国门诊机构或医院急诊科 (ED) 治疗的轻度 TBI 年发生率分别为每 10 万人中 392 例和 540 例。据报道，TBI 患者住院一年多后，残疾发生率为 37%；根据这一数字，CDC 估计全美近 2% 的人口患有与 TBI 相关的残疾。部署在阿富汗和伊拉克的士兵中，有多达 22% 可能发生创伤性脑损伤 (TBI)。最近对缺血的研究以及我们实验室未发表的工作提出了两种有趣的方法来限制损伤后细胞死亡的三种主要途径。一种方法是上调热休克蛋白 70 (HSP70)，该蛋白在不同位点结合 Apaf-1 和 AIF，从而通过阻止凋亡体（和 caspase 3）的形成并减弱 AIF 介导的作用来中和它们的促凋亡功能。行动。我们的初步数据表明，TBI 会导致损伤区内许多神经元的 HSP70 上调；那些表达 HSP 70 的神经元既没有表现出 caspase-3 激活，也没有表现出 AIF 易位。 HSP70 在脑缺血中的上调具有很强的保护作用。另一种方法是抑制 PARP-1。 PARP-1 通过释放聚 ADP 核糖 (PAR)，是神经元线粒体释放 AIF 的关键上游激活剂；最近它还被证明是小胶质细胞的关键激活剂。 TBI 后抑制 PARP-1 可强烈减弱 caspase 依赖性和非依赖性 PCD 形式以及小胶质细胞的激活，从而显着改善预后。这些策略的优点是它们的治疗窗口应该非常宽，至少 24 小时。通过结合这些不同的治疗策略，可能会实现相加或协同的保护作用。如果我们的假设得到支持，关于 TBI 治疗的概念将显着改变，治疗干预的目标人群将大大扩大。在不同物种的两个病理生物学不同模型中提出了联合治疗评估，假设跨模型和物种复制有效的治疗效果使得最终的临床转化更有可能。