Pulmonary endothelium targeted adenoviral gene therapy for the correction of mucopolysaccharidosis type I

摘要 粘多糖贮积症I型（MPS I）是一种破坏性的溶酶体储存障碍， 在10万新生儿中。MPS I的特征是α-L-艾杜糖醛酸酶（IDUA）基因突变，导致 溶酶体中糖胺聚糖的积累。虽然治疗是可行的，但它是非治愈性的， 昂贵且耗时。为此，MPS I的替代疗法受到高度追捧， 这种方法是基因治疗。虽然已经探索了几种载体系统以寻求基因治疗 对于MPS I的治疗，腺病毒载体的研究非常不足。腺病毒载体提供了几种 与其他载体相比，本发明的载体具有许多优点，例如能够复制和非复制细胞， 在不使用基因编辑技术的情况下整合到宿主基因组中， 到对以前无法到达的细胞类型进行分类的能力。因此，研究腺病毒感染的后果 MPS I介导的基因治疗可能为MPS I带来新的治疗途径。为了解 为了研究腺病毒基因治疗MPS I的结果，我将采用MPS I小鼠模型，并开发一种新的治疗方法。 腺病毒载体来递送IDUA的开放阅读框。我计划用骨髓结合基因 肽以有利地刺激内皮细胞。我假设非肝脏细胞来源可以帮助 IDUA细胞产生和肝靶向作用，以及通过基因的使用 编辑技术，可实现长期校正。我将解决这个假设，并实现 通过使用表达IDUA的髓样结合肽修饰的腺病毒载体， 使用体内测定法测定感染后血清中的IDUA水平。我也将确定实现的能力 通过使用基因编辑技术CRISPR/Cas9进行长期校正。本提案的结论 将更好地确定利用修饰的腺病毒载体作为基因治疗容器和作为 MPS I和其他遗传性单基因疾病的治疗方法。

ABSTRACT Mucopolysaccharidoses type I (MPS I) is a devastating lysosomal storage disorder that affects approximately 1 in 100,000 newborns. MPS I is characterized by mutations in the α-L-Iduronidase (IDUA) gene that lead to an accumulation of glycosaminoglycans in lysosomes. While treatment is available, it is non-curative, monetarily costly, and time-consuming. To this end, alternative therapies to MPS I have been highly sought after and one such approach is gene therapy. While several vector systems have been explored in the pursuit of a gene therapy treatment for MPS I, adenoviral vectors have been highly understudied. Adenoviral vectors offer several advantages to other vectors such as the ability to transduce both replicating and non-replicating cells, the inability to integrate into the host genome without the use of gene editing technology, and recent modifications leading to the ability to transduce previously unreachable cell types. Thus, investigating the consequences of adenoviral mediated gene therapy for MPS I could lead to new treatment avenues for MPS I. In order to understand the consequence(s) of adenoviral gene therapy for MPS I, I will employ the MPS I murine model and develop an adenoviral vector to deliver the open reading frame for IDUA. I plan to modify our vector using a myeloid-binding peptide to favorably transduce endothelial cells. I hypothesize that a non-liver cellular source can subserve the role of cellular production of IDUA and traverse liver targeting, and, through employment of gene editing technology, long-term correction can be achieved. I will address this hypothesis and achieve the goals of this proposal by employing a myeloid binding peptide modified adenoviral vector expressing IDUA and using in vivo assays to determine IDUA levels in serum post-infection. I will also determine the ability to achieve long-term correction through the use the gene editing technology CRISPR/Cas9. The findings of this proposal will better determine the feasibility of utilizing modified adenoviral vectors as gene therapy vessels and as a treatment method for MPS I and other inherited, monogenic disorders.