Somatic Stem Cells as Vectors to Deliver Biologically Active Molecules to the Inn

描述（由申请人提供）：本申请涉及广泛的挑战领域（06）使能技术和特定的挑战主题，06- dc -102：开发和验证药物和分子输送到内耳的方法。本应用的主要目标是建立和验证一种稳健、安全和持久的递送方法，利用体细胞作为载体将生物活性分子运输到内耳。体细胞干细胞，如间充质干细胞和诱导多能干细胞是自体细胞移植治疗的一个有前途的材料来源。虽然这些干细胞可用于替代内耳受损细胞，但我们最近的研究表明，移植的体细胞干细胞具有识别和迁移内耳受损螺旋神经节神经元的卓越能力。这种特性使体细胞干细胞成为一种独特的工具，可以选择性地向内耳受损细胞输送治疗剂。在Specific Aim 1中，我们将建立稳定表达高水平BDNF的间充质和诱导多能干细胞系。体外单个干细胞释放BDNF将通过创新的单细胞免疫印迹法定量评估。在特异性目标2中，释放BDNF的间充质或诱导多能干细胞将被移植到听觉神经病变动物模型的耳蜗中。这些干细胞在体内的迁移和植入将通过高分辨率的显微内窥镜成像系统和磁共振成像进行监测。此外，移植干细胞体内BDNF释放的时间变化将通过荧光活化细胞分选和Western blot分析相结合来评估。总之，这些实验将提供第一个关于使用自体体细胞干细胞将治疗剂输送到内耳的可行性的综合信息，因此具有重要的临床意义。内耳包含复杂而敏感的结构，这对治疗干预提出了重大挑战。我们建议推进一种使用患者来源的干细胞将治疗剂输送到内耳的新方法的可行性，其长期目标是使大部分患有重度感音神经性听力损失的患者受益。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06) Enabling Technologies and specific Challenge Topic, 06-DC-102: Develop and Validate Methods for Delivery of Drugs and Molecules to the Inner Ear. The primary goal of this application is to establish and validate a robust, safe and long-lasting delivery method using somatic stem cells as vectors to transport biologically active molecules into the inner ear. Somatic stem cells, such as mesenchymal stem cells and induced pluripotent stem cells represent a promising source of material for autologous cell transplantation therapies. While these stem cells can be used to replace damaged cells in the inner ear, our recent study has demonstrated that transplanted somatic stem cells exhibit remarkable abilities to identify and migrate towards damaged spiral ganglion neurons in the inner ear. This property makes somatic stem cells a unique tool to deliver therapeutic agents selectively to damaged cells in the inner ear. In Specific Aim 1, we will establish mesenchymal and induced pluripotent stem cell lines stably expressing high-level BDNF. Release of BDNF from individual stem cells in vitro will be quantitatively evaluated by innovative single cell immunoblot assays. In Specific Aim 2, mesenchymal or induced pluripotent stem cells releasing BDNF will be transplanted into the cochlea of an animal model of auditory neuropathy. Migration and engraftment of these stem cells in vivo will be monitored by a high-resolution microscopic- endoscope-based imaging system and magnetic resonance imaging. In addition, temporal changes in BDNF release from transplanted stem cells in vivo will be evaluated by a combination of fluorescence-activated cell sorting and Western blot analysis. Together, these experiments will provide the first comprehensive set of information regarding the feasibility of using autologous somatic stem cells to deliver therapeutic agents into the inner ear, and thus have significant clinical implications. The inner ear contains complex and sensitive structures, which present a significant challenge for therapeutic interventions. We propose to advance the feasibility of a novel method using patient-derived stem cells to deliver therapeutic agents to the inner ear, with the long-term goal of benefitting a large proportion of patients suffering from profound sensorineural hearing loss.