DRG engraftment of transduced mesenchymal stem cells to treat neuropathic pain

描述（由申请人提供）：神经性疼痛是常见且致残的。尽管对疼痛机制的了解有所进展，但可用的治疗方法仍然不足。本研究的总体目标是结合两种新方法，设计一种高效灵活的治疗系统。首先，针对单个（或多个）背根神经节（DRGs）的治疗将允许在节段基础上直接治疗感觉神经元，从而限制对健康感觉通路的副作用。其次，间充质干细胞（MSCs）的移植为扩大病理生理学的理解提供了一个强大的机会。具体来说，自体间充质干细胞很容易获得，它们很容易生长，它们在基因工程后具有高度分泌肽的能力，并且这些细胞不存在伦理或监管问题。初步数据表明，体外慢病毒转导在MSCs中通过许多繁殖周期产生稳定的转基因表达，因为该载体将转基因插入宿主基因组中，并且MSCs分泌的治疗肽水平比没有基因修饰的MSCs高得多。与直接向患者注射病毒载体的基因治疗方法不同，该策略避免了病毒的免疫原性和毒性作用，并允许在重新植入之前在患者体外进行可控、高效的转导，从而提高了性能和安全性。这项工作将在一系列目标中发展这种新疗法。首先，以大鼠为研究对象，研究MSCs的必要剂量、存活率和分化命运。其次，重要的解剖学和生理学观察将评估间充质干细胞移植对内源性DRG细胞的影响，以证实植入间充质干细胞本身没有有害影响。最后，概念验证实验将通过移植工程蛋白来测量大鼠神经损伤模型治疗神经性疼痛的疗效

DESCRIPTION (provided by applicant): Neuropathic pain is common and disabling. Although understanding of pain mechanisms has advanced, available treatments remain inadequate. The overall goal of the proposed research is to devise a highly effective and flexible treatment system by combining two novel approaches. First, therapy that targets a single (or several) dorsal root ganglia (DRGs) will permit direct treatment of sensory neurons on a segmental basis, thereby limiting side effects on healthy sensory pathways. Second, transplantation of mesenchymal stem cells (MSCs) provides a powerful opportunity to capitalize on expanding pathophysiological understanding. Specifically, autologous MSCs are readily available, they are easily grown, they are highly capable of secreting peptides after genetic engineering, and there are no ethical or regulatory issues with these cells. Preliminary data indicate that in vitro lentiviral transduction produces stable transgene expression in MSCs through many cycles of propagation, since this vector inserts the transgene into the host genome, and that MSCs secrete much higher levels of therapeutic peptides than they can without genetic modification. Unlike gene therapy approaches using direct viral vector injection into patients, this strategy avoids immunogenic and toxic effects of the virus, and permits controlled, highly efficient transduction outside the patient prior to reimplantation, thereby enhancing performance and safety. The proposed work will develop this new therapy in sequential Aims. First, the necessary dose of MSCs, their survival rates, and their differentiation fate will be characterized using rats as subjects. Second, important anatomical and physiological observations will evaluate effects of MSC transplantation on endogenous DRG cells to confirm that implantation of MSCs per se has no harmful effects. Finally, proof of concept experiments will measure the efficacy of treating neuropathic pain in a rat nerve-injury model, by transplantation of engineered MSCs into the DRG. The modified MSCs used in these translational trials will secrete either glial cell derived neurotrophic factor (GDNF) or interleukin 10, which have demonstrated efficacy in preventing or reversing neuropathic pain when delivered by other routes. Pain behavior and cellular effects will be compared to animals in which DRGs have been injected with MSCs that express only a reporter gene (GFP), and to untreated controls. Completion of the proposed work will establish the basis for therapeutic trials of engineered MSC as sources of these or other analgesic peptides in larger animals or human subjects.