In Vivo Editing for Hemophilia Gene Therapy

项目摘要 最近的研究表明，血友病基因治疗的特殊前景， 通过用腺相关载体（AAV）进行肝转导，长期纠正因子IX缺乏。在 尽管这是基因治疗领域历史上的分水岭事件，但目前仍存在充分利用基因治疗的障碍。 血友病基因治疗的实施。AAV对较大基因构建体的有限包装能力 实际上混淆了该载体用于因子VIII校正和载体相关毒性的用途， 值得注意的是肝脏的体内转导。我们建议开发一种新的矢量方法 解决了这些局限性。值得注意的是，我们最近的策略，重新靶向腺病毒载体（Ad）， 减轻载体相关的肝毒性和体内转导肺内皮细胞， 证明了该位点可以作为一个完全有效的平台来重建缺陷的血清蛋白。 此外，具有多个缺失的新一代广告可以被修改以并入扩大范围的 基因构建体和更大的有效载荷相关的基因治疗的血友病疾病的全谱。我们 建议利用与广告技术完全相称的CRISPR/Cas 9系统， 使用我们的肺内皮靶向Ad，以实现纠正血友病基因的稳定掺入 在肺内皮细胞内。因此，我们的新策略提供了稳定遗传校正的可能性。 以避免因子IX和因子VIII血友病的潜在载体相关毒性的方式。 在可行性证明阶段1 STTR内成功完成目标将为 将血友病矫正基因治疗应用于最广泛的患者背景，并提供基本原理 在第二阶段，我们将高度新颖的方法转化为先导化合物优化和选择。我们 研究小组在成功地将新型基因疗法从实验室转化为临床试验方面有着无与伦比的记录 战略布局我们的策略，以实现稳定表达缺陷血友病因子来源于一个 肺血管源清楚地代表了高度原创的方法。除了回避这个问题， 载体介导的肝毒性，腺病毒载体的扩大包装能力也是可行的 因子VIII和因子IX缺乏症的基因治疗。为了加快翻译我们的方法，我们 将利用来自非人灵长类动物腺病毒的腺病毒载体， 通过规避预先形成的腺病毒免疫来解决关键的人类使用障碍的载体（GAd）， 在治疗应用中显示出优异的性能特征。疗效研究概况 本文将构成这种新方法的翻译发展的基本原理，以及 建立肺内皮细胞作为提供分泌因子的来源，作为一种重要的策略， 一系列基于血清因子缺乏的遗传性疾病以及其他潜在的 治疗应用。

Project Summary The exceptional promise of gene therapy for hemophilia has been shown in recent studies that result in long term correction of factor IX deficiency, via hepatic transduction with an adeno-associated vector (AAV). In spite of this watershed event in the history of the gene therapy field, there presently exist barriers to the fullest implementation of hemophilia gene therapy. The limited packaging capacity of AAV for larger gene constructs practically confounds the use of this vector for factor VIII correction and vector-related toxicities have been noteworthy for the context of in vivo transduction of the liver. We propose to develop a novel vector approach that addresses these limitations. Of note, our recent strategies to retarget adenoviral vectors (Ad) have allowed mitigation of vector-associated liver toxicities and in vivo transduction of the pulmonary endothelium that has demonstrated that this site can serve as a fully effective platform to reconstitute deficient serum proteins. Moreover, new generation Ads with multiple deletions can be modified to incorporate an expanded range of gene constructs and larger payloads relevant to gene therapy for the full spectrum of hemophilia disorders. We propose to utilize the CRISPR/Cas 9 system which is fully commensurate with Ad technology, in conjunction with our pulmonary endothelial-targeted Ad, to achieve stable incorporation of corrective hemophilia genes within the pulmonary endothelium. Our novel strategy thereby offers the potential for stable genetic correction in a manner that circumvents potential vector-associated toxicities for both factor IX and factor VIII hemophilia. Successful completion of the aims within this proof of feasibility Phase 1 STTR will set the stage for the application of corrective gene therapy for hemophilia to the widest context of patients and provide the rationale for translational development of our highly novel approach into lead optimization and selection in Phase II. Our groups have unparalleled track records for successful bench-to-bed translation of novel gene therapy strategies. Our strategy to accomplish stable expression of deficient hemophilia factors deriving from a pulmonary vascular source clearly represents a highly original approach. In addition to circumventing the issue of vector-mediated hepatotoxicities, the expanded packaging capacity of adenovirus vectors also feasibilizes gene therapy for factor VIII, as well as factor IX deficiency disorders. To hasten translation of our approach, we will utilize adenoviral vectors derived from non-human primate adenoviruses and will specifically use gorilla Ad vectors (GAd) that address a pivotal human use barrier by circumventing preformed adenoviral immunity and are show excellent performance characteristics in therapeutic applications. The profile of efficacy studies herein will constitute the rationale for translational development of this novel approach as well as a leap to establishing the pulmonary endothelium as a source to provide secreted factors as an important strategy for the range of inherited genetic disorders based upon deficient serum factors as well as other potential therapeutic applications.