eukaryotic expression vectors resistant to transgene silencing

描述（由申请人提供）：质粒导向的基因表达现在接近功效障碍，迄今为止，这种障碍阻碍了基于质粒的人类健康应用疗法的商业化。先前的创新如电穿孔（EP）通过更有效的基因传递增加了转基因表达。在第一阶段，我们开发了有效的最小化无抗生素表达载体，并证明了通过载体设计创新可以显著改善转基因表达。我们开发了两种平台技术。”防止质粒载体载体骨架介导的转基因沉默的新组合物（抗沉默元件：ASE平台）“瞬时转染后改善质粒载体转基因表达的新载体骨架功能（瞬时表达增强剂：TEE平台）在II期中，我们假设将ASE和TEE载体与最先进的质粒传递相结合将为基因治疗创造使能的载体传递平台。在Specific Aims 1和2中，分别确定了用于骨骼肌和皮肤基因治疗的最佳ASE/TEE无抗生素载体- EP递送平台。在Specific Aim 3中，应用皮肤基因治疗平台创建了一种基于缺氧诱导因子11 （HIF-11）的治疗糖尿病足溃疡的基因药物。在Specific Aim 4中，使用角质细胞生长因子（KGF）载体-微磨皮递送组合开发了一种皮肤基因疗法来治疗皮肤老化。具体目标3和4是与约翰霍普金斯大学的伤口愈合基因治疗专家John Harmon博士和gene facellift， LLC的皮肤病学基因治疗专家Aaron Tabor博士合作完成的。本文开发的载体传递平台将进一步提高基因表达水平，使基因药物许可能够用于多种未满足的公共卫生需求。在第三阶段，糖尿病足溃疡的基因疗法和皮肤化妆品将进行临床开发。

DESCRIPTION (provided by applicant): Plasmid-directed gene expression is now near the efficacy barrier that to date has prevented commercialization of plasmid-based therapies for human health applications. Previous innovations such as electroporation (EP) increased transgene expression through more effective gene delivery. In Phase I we developed potent minimalized antibiotic-free expression vectors, and demonstrated dramatic improvements in transgene expression may be obtained through vector design innovations. Two platform technologies were developed. " Novel compositions that prevent vector-backbone mediated transgene silencing from plasmid vectors (Anti-Silencing Elements: ASE platform) " Novel vector backbone functionalities that improve transgene expression from plasmid vectors after transient transfection (transient expression enhancers: TEE platform) In Phase II we hypothesize that combining ASE and TEE vectors with state of the art plasmid delivery will create enabling vector-delivery platforms for gene therapy. In Specific Aims 1 and 2 optimal ASE/TEE antibiotic-free vector - EP delivery platforms for skeletal muscle and cutaneous gene therapy, respectively, are identified. In Specific Aim 3 the cutaneous gene therapy platform is applied to create a hypoxia-inducible factor 11 (HIF-11) based gene medicine for diabetic foot ulcer treatment. In Specific Aim 4 a dermatological gene therapy to treat skin aging is developed using a keratinocyte growth factor (KGF) vector- microdermabrasion delivery combination. Specific Aims 3 and 4 are performed in collaboration with wound healing gene therapy expert Dr. John Harmon at Johns Hopkins University and dermatology gene therapy expert Dr Aaron Tabor at Gene Facelift, LLC. The vector-delivery platforms developed herein will further improve gene expression to levels that will enable gene medicine licensure for multiple applications for unmet public health needs. In Phase III the gene therapies for diabetic foot ulcers and skin cosmetics will undergo clinical development. PUBLIC HEALTH RELEVANCE: The objective of this proposal is to validate a novel antibiotic-free non-viral gene therapy platform, and as such is responsive to NIGMS SBIR high-priority area of interest in development of improved vectors for gene transfer. The vectors contain transient expression enhancers that improve transgene expression level and duration after gene delivery to skin or muscle. The platform will be applied to create gene therapy products to treat diabetic neuropathic foot ulcers and skin aging.