Biodegradable Mucus Penetrating DNA Nanoparticle for Gene Therapy of CF

 描述：囊性纤维化(CF)基因治疗可能使所有囊性纤维化患者受益，而不考虑导致疾病状态的特定基因突变。然而，使用病毒和非病毒载体的基因治疗试验，由于不能，迄今的结果令人失望。 克服生物屏障，包括化脓性痰凝胶层、纤毛层和上皮表面。特别是，我们先前发现，几种临床和临床前使用的病毒和非病毒基因载体被困在人的CF痰中，这阻止了它们到达并将其治疗基因货物运送到下面的呼吸道上皮细胞。我们最近开发了两种给药策略：(I)超小DNA载药纳米粒，称为粘液穿透DNA纳米粒(DNA-MPP)，它可以有效地穿透人的CF痰和(Ii)轻度低渗水载体，它通过渗透驱动的液体吸收进一步促进DNA-MPP通过痰凝胶层和纤毛层的渗透，并通过调节体积减少机制增强呼吸道上皮对DNA-MPP的摄取。我们发现，这两种方法的结合导致了有效的基因转移到体内正常小鼠的呼吸道上皮细胞和体外培养的原代人呼吸道细胞中。在这里，我们建议在相关的体外和体内环境中进一步开发和验证我们的组合方法。为此，我们将评估CFTR基因转移到生长在气液界面的CFF人原代呼吸道上皮细胞和CFR小鼠模型的肺内，以明确其CFR样肺部疾病的特征。我们还将确定我们的递送策略是否允许在单次和重复剂量下持续表达转基因，而不会引发急性和慢性毒性。如果我们成功了，我们将在猕猴身上测试我们的策略 并有可能在未来进行临床研究。

 DESCRIPTION: Cystic fibrosis (CF) gene therapy can potentially benefit all CF patients, regardless of the specific gene mutation leading to the disease state. However, gene therapy trials, using viral and non-viral vectors, have had disappointing outcomes to date due to inability to overcome biological barriers, including the purulent sputum gel layer, periciliary layer and epithelial surface. In particular, we previously found that several clinically and preclinically teted viral and non-viral gene vectors are trapped in human CF sputum, which prevents them from reaching and delivering their therapeutic gene cargo to the underlying airway epithelium. We recently developed two delivery strategies: (i) ultra-small DNA-loaded nanoparticles, called mucus-penetrating DNA nanoparticles (DNA-MPP) that efficiently penetrate human CF sputum and (ii) mildly hypotonic aqueous vehicles, which further improves DNA-MPP penetration through the sputum gel layer and periciliary layer via osmosis-driven fluid absorption, and enhances DNA-MPP uptake by the airway epithelium via the regulatory volume decrease mechanism. We found that the combination of these two approaches led to efficient gene transfer to the airway epithelium of normal mice in vivo and primary human airway cells in vitro. Here we propose to further develop and validate our combined approach in relevant in vitro and in vivo settings. To this end, we will evaluate CFTR gene transfer to CF human primary airway epithelium grown at air-liquid interface and in the lung airways of CF mouse model thoroughly characterized for its CF-like lung diseases. We will also determine whether our delivery strategies allow persistent transgene expression upon a single and repeated dosing without eliciting acute and chronic toxicity. If we are successful, we will test our strategies in macaques and potentially clinical studies in the future.