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)使能技术和具体的挑战主题，06-DC-102：开发和验证将药物和分子输送到内耳的方法。这项应用的主要目标是建立和验证一种强大、安全和持久的给药方法，使用体细胞作为载体将生物活性分子输送到内耳。体细胞干细胞，如间充质干细胞和诱导的多能干细胞，是自体细胞移植治疗的一个有前途的材料来源。虽然这些干细胞可以用来替代内耳中受损的细胞，但我们最近的研究表明，移植的体细胞干细胞具有显著的识别和迁移内耳受损螺旋神经节神经元的能力。这一特性使体细胞干细胞成为一种独特的工具，可以选择性地将治疗剂输送到内耳受损的细胞。在具体目标1中，我们将建立稳定表达高水平BDNF的间充质干细胞系和诱导多能干细胞系。将通过创新的单细胞免疫印迹分析来定量评估单个干细胞在体外释放BDNF的情况。在特定的目标2中，间充质干细胞或诱导的释放BDNF的多能干细胞将被移植到听神经病动物模型的耳蜗内。这些干细胞在体内的迁移和植入将通过基于高分辨率显微内窥镜的成像系统和磁共振成像进行监测。此外，体内移植干细胞释放BDNF的时间变化将通过荧光激活细胞分类和Western印迹分析相结合的方法进行评估。总之，这些实验将提供第一组关于使用自体体细胞将治疗剂输送到内耳的可行性的全面信息，因此具有重要的临床意义。内耳包含复杂而敏感的结构，这给治疗干预带来了巨大的挑战。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，长期目标是使大部分患有严重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.