Enhanced intratympanic delivery of therapeutics to treat and prevent hearing loss using nanovesicles in the porcine model

在猪模型中使用纳米囊泡增强治疗剂的鼓室内递送以治疗和预防听力损失

• 批准号: 10525059
• 负责人: Adele Moatti
• 金额: $ 10.23万
• 依托单位: NORTH CAROLINA STATE UNIVERSITY RALEIGH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10525059
• 关键词: Address; Affect; Amino Acid Sequence; Animal Model; Animals; Biological Models; Cell membrane; Cells; Clinical Treatment; Clinical Trials; Cochlea; Data; Diffuse; Drug Screening; Ear; Encapsulated; Excision; FDA approved; Family suidae; Genes; Genetic; Goals; Gold; Hearing; Human; In Vitro; Individual; Injections; Labyrinth; Liposomes; Measures; Membrane; Mesenchymal Stem Cells; Methods; Mission; Modeling; Natural regeneration; Operative Surgical Procedures; Outcome; Pathology; Perilymph; Permeability; Pharmaceutical Preparations; Phase; Physiology; Procedures; Proteins; Public Health; Reporting; Research; Research Personnel; Risk; Rodent; Sampling; System; Testing; Therapeutic; Thick; Time; Translating; Trauma; Tympanic membrane; United States National Institutes of Health; Work; deafness; disability; efficacy testing; exosome; extracellular; extracellular vesicles; gene therapy; hair cell regeneration; hearing impairment; hearing loss treatment; human tissue; improved; in vivo; membrane model; nanocarrier; nanovesicle; neuron regeneration; neurotrophic factor; novel; novel therapeutics; porcine model; pre-clinical; prevent hearing loss; regenerative therapy; round window; small molecule; stem cell exosomes; success; tool; uptake

PROJECT SUMMARY My long-term goal is to help develop and deliver regeneration therapeutics useful for the clinical treatment of hearing loss. Unfortunately, at this time there is no high throughput, non-destructive method of therapeutic delivery into the cochlea/inner ear. The objectives of this proposal, the next step toward the attainment of this long-term goal, are to use a relevant large animal model (the pig) to i) develop an ex-vivo model system, which mimics human round window membrane (RWM), to track the passage of therapy-related substances through the RWM, and ii) create a method to enhance the intratympanic delivery of therapeutics into the inner ear using nanovesicles including extracellular vesicles (exosomes). The central hypothesis, supported by our preliminary data, is that an ex-vivo porcine RWM model can be used to determine optimal conditions for the transport of exosomes (or other nanovesicles) to deliver cargo through the RWM into the inner ear, thereby facilitating the high-efficiency delivery of therapeutics. The rationale for this proposal is that its successful completion is likely to offer a framework whereby a new pool of therapeutics can be tested for non-surgical delivery into the inner ear, in a large animal model that has the preclinical advantage over rodents in terms of size, physiology, and genetic similarity (amino-acid sequences of common deafness genes) to humans. The following specific aims will be pursued during K99 (aim 1 and 2) and R00 phase: Aim 1) To further validate the ex-vivo porcine RWM model to demonstrate drug permeability equivalent to that reported for human tissue; Aim 2) To identify and evaluate exosomes and other nanovesicles enhancing cargo transport across porcine RWM in-vitro; Aim 3) To evaluate in-vivo transport across porcine RWM by nanovesicles. Under the first aim, we will measure the permeability of therapeutics with known delivery efficiencies and some promising therapy related materials using a preliminary viability-verified ex-vivo RWM model and compare those with values reported for human tissue. For the second aim, nanovesicles including: RWM exosomes (successfully isolated and characterized as a preliminary result), mesenchymal stem cell exosomes (gold standard), and liposomes (FDA approved), that are loaded with promising therapeutics will be evaluated for the efficiency of transport using the ex-vivo RWM model. Finally, for the last aim, once the parameters associated with optimal transport through the RWM are established with the ex-vivo model, we will inject therapeutic-loaded nanovesicles through the porcine tympanic membrane and measure their passage across RWM and uptake by cochlear cells in vivo. Upon completion of the K99, the expected outcomes are 1. Availability of a safe and translatable platform to test transport of therapeutics into the inner ear, and 2. Data on the efficiency of nanovesicles as novel nonsurgical transport of promising therapeutics to the ear. These results are expected to have a positive impact because they could improve drug screening for delivery and is likely to boost delivering novel regenerative therapeutics to treat and prevent hearing loss.

项目总结 我的长期目标是帮助开发和提供对临床治疗有用的再生疗法 听力损失。遗憾的是，目前还没有高通量、非破坏性的治疗方法 传输到耳蜗处/内耳。这项提案的目标，实现这一目标的下一步 长期目标，是利用相关的大型动物模型(猪)来开发一种体外模型系统，它 模拟人类圆形窗膜(RWM)，追踪治疗相关物质通过 RWM，以及ii)创造了一种方法，以增强治疗药物在内耳中的输送 纳米囊泡，包括胞外囊泡(外体)。核心假设，得到了我们初步的支持 数据表明，体外猪RWM模型可用于确定药物转运的最佳条件。 外体(或其他纳米囊泡)通过RWM将货物输送到内耳，从而促进 高效率地提供治疗药物。这项提议的理由是，它很有可能成功完成 提供一个框架，以便可以测试新的治疗方法池是否可以通过非手术方式进入体内 耳，在一个大型动物模型中，在大小、生理和 遗传相似性(常见耳聋基因的氨基酸序列)与人类。以下是具体目标 将在K99(目标1和2)和R00阶段：目标1)期间进行，以进一步验证体外猪RWM 证明药物对人体组织的渗透性等同于已报道的人体组织渗透性的模型；目的2)识别和 评估外周小体和其他纳米囊泡在体外促进通过猪RWM的货物运输；目标3) 评价纳米微囊在猪RWM中的体内转运。在第一个目标下，我们将衡量 已知给药效率的治疗药物的渗透性和一些有前景的治疗相关材料 一个初步的活性验证的体外RWM模型，并将其与报告的人体组织值进行比较。 对于第二个目的，纳米囊泡包括：RWM外切体(成功分离并表征为 初步结果)、间充质干细胞外切体(黄金标准)和脂质体(FDA批准)，它们是 将使用体外RWM模型评估装载有前景的治疗药物的运输效率。 最后，为了最后的目的，一旦建立了与通过RWM的最佳传输相关联的参数 在体外模型中，我们将通过猪鼓膜注射治疗负载的纳米微囊 并测量它们通过RWM的途径和体内耳蜗细胞的摄取。K99号公路建成后， 预期结果是1.提供一个安全和可翻译的平台，以测试治疗药物进入 内耳，以及2.纳米囊泡作为有前景的治疗药物的新的非手术传输的效率的数据 送到耳朵里。这些结果预计将产生积极影响，因为它们可以改善药物筛选 并可能促进提供治疗和预防听力损失的新型再生疗法。