Ionizable Lipid Nanoparticles for Fetal Lung Targeting

项目总结 先天性肺部疾病，如遗传性表面活性物质蛋白综合征、囊性纤维化和 α-1抗胰蛋白酶缺乏症是儿科发病率和死亡率的重要来源。 合并呼吸衰竭的新生儿肺部疾病的治疗选择 仅限于姑息治疗或儿童肺移植。因此，有明确的临床需求。 为早期纠正先天性肺部疾病提供新的治疗方法，以减少儿童 发病率和死亡率。CRISPR-Cas9等基因编辑技术的最新进展 系统，释放了纠正致病突变的可能性，从而治疗先天性 从源头上解决问题。在子宫中进行基因编辑提供了逆转的额外好处 当肺功能转变为出生后生活之前的遗传异常 必要的，并利用胎儿的正常发育特性来提高效率 更正。传统上，病毒载体被用于动物子宫基因治疗的研究。 模特们。尽管这些研究令人鼓舞，但发现了替代的、潜在的更安全的 送货车辆将把这一领域推向临床翻译。因此，这项提案旨在 研究可电离脂质纳米粒(LNPs)的潜力，这是一种有前途的非病毒递送材料 用于将核酸输送到小鼠胎肺的平台。优化的胎肺脂质纳米粒 (Flo-LNPs)将通过多阶段优化方案生成。在目标1中，一个多样化的 将筛选24个可电离类脂结构的文库，以确定最好的可电离类脂 将信使核糖核酸输送到胎儿肺。在目标2中，将使用以下设计优化LNP配方 CRISPR-CAS9系统最小毒性和最大投放量的实验方案 小鼠精确切割肺切片。在目标3中，将通过以下方式修改优化的LNP配方 抗体结合，并在胎鼠肺中进行细胞特异性靶向测试。归根结底，这 提案--作为新闻部赞助者之间的跨学科项目进行 生物工程，佩雷尔曼医学院和费城儿童医院 宾夕法尼亚大学-将允许开发一种新型的LNP交付平台 可应用于随后的工作，为先天性肺部疾病提供宫内基因治疗。

PROJECT SUMMARY Congenital lung diseases, such as inherited surfactant protein syndromes, cystic fibrosis, and alpha-1 antitrypsin deficiency, are a significant source of pediatric morbidity and mortality. Treatment options for neonatal patients with lung disorders that present with respiratory failure are limited to palliative care or pediatric lung transplant. As such, there is a clear clinical demand for new therapies that allow for early correction of congenital lung diseases to reduce pediatric morbidity and mortality. Recent advances in gene editing technologies, such as CRISPR-Cas9 systems, have unlocked the potential to correct pathogenic mutations and thereby treat congenital disorders at their source. Performing gene editing in utero offers the added benefits of reversing genetic abnormalities prior to the transition to postnatal life, when pulmonary function becomes essential, and harnessing normal developmental properties of the fetus for more efficient correction. Traditionally, viral vectors have been used to study in utero gene therapy in animal models. Although these studies are encouraging, discovery of alternative, potentially safer, delivery vehicles will advance the field toward clinical translation. Thus, this proposal aims to investigate the potential of ionizable lipid nanoparticles (LNPs), a promising non-viral delivery platform, for nucleic acid delivery to the mouse fetal lung. Fetal lung optimized lipid nanoparticles (FLO-LNPs) will be generated through a multi-stage optimization scheme. In Aim 1, a diverse library of 24 ionizable lipid structures will be screened to identify the ionizable lipid that best delivers mRNA to the fetal lung. In Aim 2, LNP formulations will be optimized using a Design of Experiments scheme for minimal toxicity and maximal delivery of a CRISPR-Cas9 systems in mouse precision cut lung slices. In Aim 3, the optimized LNP formulation will be modified via antibody conjugation and tested for cell-specific targeting in the fetal mouse lung. Ultimately, this proposal – conducted as an interdisciplinary project between sponsors in the Department of Bioengineering, Perelman School of Medicine, and Children’s Hospital of Philadelphia at the University of Pennsylvania – will allow for the development of a novel LNP delivery platform that can be applied in subsequent work to deliver in utero gene therapies for congenital lung disease.