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

摘要 粘多糖I型(MPS I)是一种破坏性的溶酶体储存障碍，影响约1 在10万名新生儿中。MPS I的特征是α-L-艾杜糖苷酶基因突变导致 糖胺多聚糖在溶酶体中的积累。虽然治疗是可用的，但从金钱上讲，它是不能治愈的。 成本高、耗时长。为此，MPS的替代疗法一直备受追捧，其中一种 这种方法就是基因疗法。虽然已经探索了几种载体系统来追求基因疗法 对于MPS I的治疗，腺病毒载体的研究一直很少。腺病毒载体提供了几个 其他载体的优点，如转导复制和非复制细胞的能力，不能 在不使用基因编辑技术的情况下整合到宿主基因组中，最近的修改导致 涉及到转换以前无法到达的小区类型的能力。因此，调查腺病毒的后果 介导性MPS I基因治疗可能为MPS I开辟新的治疗途径。 腺病毒基因治疗MPS I的结果(S)，我将采用MPS I小鼠模型，并建立 腺病毒载体为IDUA提供开放阅读框架。我计划用髓系结合来修饰我们的载体 有利于转导内皮细胞的多肽。我假设非肝细胞来源可以替代 IDUA在细胞生产中的作用和肝靶向性，并通过利用基因 编辑技术，可实现长期改版。我将解决这一假设，并实现 通过使用髓系结合肽修饰的腺病毒载体表达IDUA和 用体内测定法测定感染后血清中IDUA水平。我还将确定是否有能力实现 通过使用CRISPR/CAS9基因编辑技术进行长期矫正。这项提案的结果是 将更好地确定利用改良腺病毒载体作为基因治疗血管和作为 治疗MPS I和其他遗传性、单基因疾病的方法。

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.