Combined Nano and Cell therapy for the Treatment of TBI

 描述（由申请人提供）： 我们研究计划的长期目标是开发安全有效的疗法来治疗创伤性脑损伤 (TBI)，这是我们在战斗中的军事人员面临的一个主要健康问题，据估计，伊拉克自由行动和持久自由行动中有 150,000-300,000 名军事人员遭受了一定程度的 TBI。据估计，仅在美国，各种原因导致的 TBI 患病率就达到 2.5 至 370 万人。目前，尚无药物可用于治疗 TBI。虽然TBI疾病的范围从轻度到重度不等，但中度TBI在临床上最为相关，如果不及时治疗，通常会进展为难以治疗的重度TBI。因此，确实需要发现中度TBI的新治疗剂。包括间充质干细胞 (MSC) 在内的成体干细胞疗法正在研究中，然而，由于使用不同的模型和技术、输送途径的不确定性、植入的可变性以及不良事件的可能性，对于这种 MSC 疗法的可行性仍缺乏共识。因此，发现和开发新的治疗靶点仍然是一个未满足的主要需求。我们做出了惊人的发现，趋化因子（C-C 基序）配体 20 (CCL20) (MIP3α) 的表达对于啮齿类动物 LFP-TBI（横向流体冲击 (LFP) 损伤模型）后的炎症至关重要，这表明 CCL20 作为潜在的 TBI 药物靶点。我们的数据表明，TBI 的急性期涉及 CCL20 驱动的炎症细胞从外周涌入，这些细胞似乎在随后的神经炎症和神经变性中发挥作用。为了开发一种基于 CCL20 的 TBI 纳米基因药物，我们合成了多功能壳聚糖-聚乙烯亚胺 (PEI) 磁性胶束 (CPMM)，将药物和基因输送到受伤的大脑。此外，我们发现鼻内递送的 MSC 在 TBI 后很容易迁移到大脑和脾脏，并减少受伤大脑中的反应性星形胶质细胞增生。这些结果使我们推测，组合纳米细胞疗法包括在TBI后1-2天鼻内/静脉内施用CPMM以递送针对CCL20的小干扰RNA（siRNA），然后在TBI后3-10天进行MSC治疗，将分别改善急性炎症和继发性神经元损伤，从而为中度TBI提供有效的治疗。提出了三个具体目标来检验这一假设。在目标#1中，我们将合成并表征针对 CCL20 信号传导的 TRIC（靶向活性氧诱导型 CPMM）纳米颗粒，并检查 CCL20 抑制对 LFP 诱导的 TBI 病理的影响。在目标#2中，我们将评估为 MSC 递送提供更好的治疗窗口的递送途径，以及 MSC 递送在修复 LFP 诱导的中度 TBI 后大鼠大脑神经功能缺陷中的功效。在目标 3 中，我们计划评估两步纳米细胞治疗方法对中度 TBI 的治疗效果。我们将比较靶向 CCL20/CCR6 的 TRIC NP 和 MSC 治疗在改善中度 TBI 后的缺陷方面的功效，并确定作用机制。这种高度跨学科和创新的两步疗法方法涉及携带 CCL20 信号抑制剂的 NP 和 hMSC 疗法，用于治疗 LFP 诱导的中度 TBI，具有高度创新性。我们预测，拟议的研究将加深我们对 TRIC-siCCL20 在 TBI 中的潜力和作用的理解。所有必要的试剂、方法和合作都已到位，我们的研究团队已经做好了进行拟议研究的独特准备。

 DESCRIPTION (provided by applicant): The long-term goal of our research program is to develop safe and effective therapies for traumatic brain injury (TBI), which is a major health issue with our military personnel in combat, with estimates of 150,000-300,000 military personnel from Operation Iraqi Freedom and Operation Enduring Freedom suffering some level of TBI. The prevalence of TBI from all causes has been estimated to be 2.5 to 3.7 millions in the USA alone. Currently, there are no drugs available to treat TBI. While TBI illness ranges from mild to severe, moderate TBI is clinically most relevant, and if left untreated it often progresses to severe TBI, which is difficult to treat Hence, there is a genuine need to discover new therapeutic agents for moderate TBI. Adult stem cell therapy including mesenchymal stem cells (MSCs), is under investigation, however, there remains a lack of consensus about the viability of this MSC therapy due to use of different models and techniques, uncertainty about routes of delivery, variable engraftment and the potential for adverse events. Thus, discovering and developing new therapeutic targets remains a major unmet need. We made the striking discovery that expression of chemokine (C-C motif) ligand 20 (CCL20) (MIP3α) is pivotal to inflammation following LFP-TBI (a lateral fluid percussion (LFP) injury model) in rodents, implicating CCL20 as a potential TBI drug target. Our data demonstrates that the acute phase of TBI involves an influx of CCL20-driven inflammatory cells from the periphery that appear to play a role in subsequent neuroinflammation and neurodegeneration. Toward developing a CCL20-based nanogene drug for TBI, we have synthesized multifunctional chitosan- polyethyleneimine (PEI) magnetic micelles (CPMMs) to deliver drugs and genes to the injured brain. In addition, we have found that intranasally delivered MSCs readily migrate to brain and spleen post TBI and reduce reactive astrogliosis in the injured brain. These results have led us to hypothesize that a combinatorial nano-cell therapy comprising intranasal/intravenous administration of CPMMs 1-2 days after TBI to deliver small interfering RNAs (siRNAs) against CCL20 followed by MSC therapy 3-10 days after TBI would respectively, ameliorate acute inflammation and secondary neuronal injury thus providing an effective treatment for moderate TBI. Three specific aims are proposed to test this hypothesis. In Aim #1, we will synthesize and characterize TRIC (targeted, reactive oxygen species-inducible CPMM) nanoparticles targeting CCL20 signaling and examine the effects of CCL20 inhibition on LFP-induced TBI pathology. In Aim #2, we will evaluate the route of delivery that provides a better therapeutic window for MSC delivery and the efficacy of MSC delivery in repairing neurological function deficits in the rat brain after LFP-induced moderate TBI. In Aim #3, we plan to evaluate therapeutic efficacy of a two-step nano-cell treatment approach for moderate TBI. We will compare the efficacy of TRIC NPs targeting CCL20/CCR6 followed by MSC therapy in ameliorating deficits following moderate TBI and determine mechanisms of action. This highly interdisciplinary and innovative approach of a two-step therapy involving NPs carrying the inhibitor(s) of CCL20 signaling and hMSC therapy for treatment of LFP-induced moderate TBI is highly innovative. We predict that the proposed research will increase our understanding of the potential and role of TRIC-siCCL20 for TBI. All the necessary reagents, methods and collaborations are in place and our investigative team is uniquely poised to conduct the proposed studies.