Gene targeting with AAV vectors for the treatment of Osteogenesis Imperfecta

描述（由申请人提供）：本提案描述了一个为期五年的培训计划，以发展医学遗传学和基因治疗的学术生涯。该计划将进一步推进研究人员在开发用于治疗成骨不全的AAV介导的基因靶向策略方面的研究。研究人员将由大卫罗素博士指导，他在开发用于基因靶向的腺相关病毒（AAV）载体系统方面的成就得到了广泛认可。成骨不全症（01）是一组遗传性胶原蛋白疾病，其特征是骨脆性，临床表现从轻度骨折增加到严重骨畸形和死亡不等。导致OI的突变位于COL 1A 1和COL 1A 2基因中。目前的治疗选择是有限的，并没有减轻在OI中看到的并发症。大多数基因治疗系统涉及基因添加的过程，其中启动子和基因被递送到细胞中并随机整合。相比之下，基因靶向使用同源序列以位点特异性方式改变细胞的内源基因。通过这种方法，突变的基因可以被“敲除”或校正。基因打靶的主要优点是基因可以在染色体上的适当位置进行修饰，而不会破坏细胞的基因调控。Russell博士先前的工作已经证明，AAV介导的基因靶向载体可以靶向COL 1A 1基因并破坏显性阴性突变型前胶原的产生。结果表明，突变蛋白的缺失改善了胶原原纤维的加工性、稳定性和结构。该提议的目的是开发AAV基因靶向载体，其将靶向COL 1A 2基因，减少随机整合，并靶向具有Ol的多个个体。具体目标包括：1）开发一种腺相关病毒基因靶向载体，以靶向Ol中的COL 1A 2基因; 2）开发一种改进的腺相关病毒基因靶向载体，以减少随机整合体的回收;和3）确定人类变异对基因靶向的影响。该项目的完成和在华盛顿大学接受的培训将使主要研究者成为基因治疗领域的主要研究人员。

DESCRIPTION (provided by applicant): This proposal describes a five-year training program to develop an academic career in Medical Genetics and gene therapy. The program will further advance the investigator's research in the development of an AAV-mediated gene targeting strategy for the treatment of Osteogenesis Imperfecta. The investigator will be mentored by Dr. David Russell, well recognized for his achievements in developing the adeno-associated virus (AAV) vector system for gene targeting. Osteogenesis Imperfecta (01) is a group of inherited collagen disorders characterized by bone fragility with clinical manifestations varying from a mild increase in fractures to severe bone deformities and death. Mutations that cause Ol are located in the COL1A1 and COL1A2 genes. Current treatment options are limited and do not alleviate the complications seen in Ol. Most gene therapy systems involve the process of gene addition, where a promoter and a gene are delivered into a cell and integrate randomly. In contrast, gene targeting uses homologous sequences to alter the cell's endogenous genes in a site specific manner. By this method mutated genes can be either "knocked-out" or corrected. The major advantage of gene targeting is that genes can be modified at their proper location on the chromosme without disrupting cellular gene regulation. Previous work by Dr. Russell has demonstrated that an AAV-mediated gene targeting vector can target the COL1A1 gene and disrupt dominant negative mutant procollagen production. It was shown that the loss of the mutant protein improved the processing, stability and structure of the collagen fibril. The objective of this proposal is to develop an AAV gene targeting vector that will target the COL1A2 gene, reduce random integration, and target multiple individuals with Ol. The specific aims include: 1) Develop an AAV gene-targeting vector to target the COL1A2 gene in Ol; 2) Develop an improved AAV gene-targeting vector to reduce the recovery of random integrants; and 3) Determine the effects of human variation on gene targeting. The completion of this project and the training received at the University of Washington will prepare the principle investigator to be a leading researcher in gene therapy.