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Histone targeted non-viral gene delivery to enhance bone repair

组蛋白靶向非病毒基因传递以增强骨修复

基本信息

批准号：
9229553
负责人：
Millicent O Sullivan
金额：
$ 34.98万
依托单位：
UNIVERSITY OF DELAWARE
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-05-01 至 2019-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9229553
关键词：
Address Area BMP2 gene Binding Biocompatible Materials Biology Bone Injury Bone Regeneration Bone Transplantation Cations Cell Differentiation process Cells Chemistry Chromatin Clinical Clinical Research Complex Coupled DNA DNA Binding Defect Development Exhibits Fostering Fracture Gene Delivery Gene Expression Gene Targeting Gene Transfer Genes Genetic Transcription Goals Growth Factor Growth Factor Gene Health Heterotopic Ossification Histones Image Image Analysis Implant In Situ In Vitro Individual Knowledge Letters Link Mediating Mediation Mesenchymal Differentiation Mesenchymal Stem Cells Methods Modeling Monitor Mus Mutagenesis Natural regeneration Nature Nuclear Operative Surgical Procedures Orthopedics Osteogenesis Outcome Pathway interactions Peptides Physiologic Ossification Post-Translational Protein Processing Process Production Productivity Proteins Publishing Rattus Regulator Genes Reporting Role Safety Speed Structure System Tail Testing Therapeutic Tissue Engineering Transfection Translations Viral Viral Genes base biomaterial compatibility bone bone healing bone morphogenetic protein 2 clinical efficacy combinatorial controlled release cost design economic cost gene delivery system gene product gene therapy healing imaging potential immunogenicity improved in vivo insight interest mimetics nanoGold nanoparticle new technology non-viral gene delivery novel osteogenic plasmid DNA public health relevance release factor repaired scaffold self assembly spatiotemporal success tissue repair trafficking uptake viral gene delivery

项目摘要

DESCRIPTION (provided by applicant): Large segmental bone defects are a persistent clinical challenge that poses significant economic costs as well as costs to individual health and societal productivity. Limitations in bone grafting emphasize the need for alternative strategies. Accordingly, a myriad of therapeutic approaches have been explored to address this health problem, including the direct delivery of growth factors and delivery of viral growth factor gene therapies. While these approaches have improved healing outcomes, they have not been widely employed clinically, owing at least in part to limited protein stability (direct delivery) and safey concerns (viral gene therapies). Non-viral strategies to induce efficient, cell-mediated production of growth factors would offer a provocative approach to overcome these difficulties. We propose to address these challenges through the development of new, histone-targeted gene transfer scaffolds with tunable DNA binding and controllable gene delivery. The display of histone motifs on nanoparticle scaffolds (e.g. nanogold) will capitalize on our preliminary studies proving that post-translationally modified histone tails promote nuclear accumulation, DNA release, transcription, and enhanced transfection by non-viral vehicles. Additionally, our approach builds on established advantages of nanogold, including imaging potential, biocompatibility, functionalizability, and cell entry capacity. The novel presentation of histone tails on nanogold should mimic the native, multifaceted display and functionality of these sequences on the histone octamer. Hence, these new scaffolds should provide additional important, yet unexplored, benefits for controlling and understanding gene packaging, trafficking, and release through enhanced utilization of native gene transfer and trafficking pathways. Accordingly, the goal of this proposal is to create and optimize multifunctional "designer" histones to induce efficient gene transfer, and ultimately, enable improved tissue repair for orthopedics and other applications. We will produce nanogold-plasmid DNA (pDNA) assemblies, and will determine whether the multivalent presentation of pDNA-binding residues and histone peptides enhances pDNA-binding stability and improves transfection. We will capitalize on traditional as well as nanogold-specific imaging analyses to elucidate key steps in non-viral gene transfer, and will clearly link enhanced gene transfer efficiency to improved osteogenic potential of growth factors such as bone morphogenetic protein-2 (BMP-2). Finally, we will use murine and rat orthopedic models to test the ability of the BMP-2 gene product to enhance bone formation and increase the speed and efficacy of defect repair. Our approaches will not only elucidate mechanistic details about the trafficking of histone-associated genes, but will also ultimately will be useful as a general biomaterials platform applicable to bone repair, implant functionalization, and tissue engineering.

描述(由申请人提供)：大型节段性骨缺损是一项长期的临床挑战，不仅对个人健康和社会生产力造成巨大的经济成本。骨移植的局限性强调了替代策略的必要性。因此，已经探索了无数的治疗方法来解决这一健康问题，包括直接输送生长因子和输送病毒生长因子基因疗法。虽然这些方法改善了愈合结果，但它们尚未广泛应用于临床，至少部分原因是蛋白质稳定性有限(直接输送)和安全性问题(病毒基因疗法)。非病毒策略诱导高效的细胞介导的生产增长因素的变化将为克服这些困难提供一种具有挑衅性的方法。我们建议通过开发具有可调节的DNA结合和可控制的基因输送的新的、组蛋白靶向的基因转移支架来应对这些挑战。在纳米颗粒支架(例如纳米金)上展示组蛋白基序将利用我们的初步研究证明翻译后修饰的组蛋白尾巴促进非病毒载体的核积累、DNA释放、转录和增强转染。此外，我们的方法建立在纳米金的既定优势之上，包括成像潜力、生物兼容性、功能化能力和细胞进入能力。组蛋白尾巴在纳米金上的新颖呈现应该模仿这些序列在组蛋白八聚体上的天然、多方面的显示和功能。因此，这些新的支架应该通过加强利用本地基因转移和贩运途径，为控制和理解基因包装、贩运和释放提供额外的重要但尚未探索的好处。因此，这项建议的目标是创造和优化多功能“设计者”组蛋白，以诱导有效的基因转移，并最终实现骨科和其他应用的改进组织修复。我们将生产纳米金质粒DNA(PDNA)组装，并将确定PDNA结合残基和组蛋白多肽的多价呈现是否提高了PDNA结合的稳定性和改善了转染率。我们将利用传统的和纳米金特异的成像分析来阐明非病毒基因转移的关键步骤，并将明确地将提高的基因转移效率与改善骨形态发生蛋白-2(BMP-2)等生长因子的成骨潜力联系起来。最后，我们将使用小鼠和大鼠骨科模型来测试BMP-2基因产物促进骨形成和提高缺损修复的速度和效果的能力。我们的方法不仅将阐明组蛋白相关基因运输的机制细节，而且最终将作为适用于骨修复、种植体功能化和组织工程的通用生物材料平台。