Biodegradable Mucus Penetrating DNA Nanoparticle for Gene Therapy of CF

 产品说明：囊性纤维化（CF）基因治疗可能使所有CF患者受益，无论导致疾病状态的特定基因突变如何。然而，使用病毒和非病毒载体的基因治疗试验迄今为止由于无法实现基因治疗，结果令人失望。 克服生物屏障，包括脓痰凝胶层、纤毛周层和上皮表面。特别是，我们以前发现，几个临床和临床前测试的病毒和非病毒基因载体被困在人CF痰液中，这阻止了它们到达并将其治疗基因货物递送到下面的气道上皮。我们最近制定了两项交付战略：（i）负载DNA的超小纳米颗粒，称为粘液穿透DNA纳米颗粒（DNA-MPP），其有效地穿透人CF痰液，和（ii）轻度低渗水性载体，其通过痰液驱动的流体吸收进一步改善DNA-MPP穿透痰液凝胶层和纤毛周层，并通过调节性容积减少机制增强气道上皮细胞对DNA-MPP的摄取。我们发现，这两种方法的组合导致有效的基因转移到正常小鼠的气道上皮细胞在体内和原代人气道细胞在体外。在这里，我们建议进一步开发和验证我们的组合方法在相关的体外和体内环境。为此，我们将评估CFTR基因转移到在气液界面生长的CF人原代气道上皮细胞和CF小鼠模型的肺气道中，其特征在于其CF样肺病。我们还将确定我们的递送策略是否允许在单次和重复给药后持续的转基因表达而不引起急性和慢性毒性。如果我们成功了，我们将在猕猴身上测试我们的策略 以及将来可能的临床研究。

 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.