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

• 批准号: 8613949
• 负责人: Millicent O Sullivan
• 金额: $ 35.3万
• 依托单位: UNIVERSITY OF DELAWARE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8613949
• 关键词: Address; Area; BMP2 gene; Binding; Biocompatible Materials; Biology; Bone Injury; Bone Regeneration; Bone Transplantation; Cell physiology; Cells; Chemistry; Chromatin; Clinical; Clinical Research; Complex; Coupled; DNA; DNA Binding; Defect; Development; Exhibits; Fostering; Fracture; Gene Delivery; Gene Expression; Gene Transfer; Genes; Genetic Transcription; Goals; Growth Factor; Growth Factor Gene; Healed; Health; Heterotopic Ossification; Histones; Image; Image Analysis; Implant; In Situ; In Vitro; Individual; Knowledge; Letters; Link; Mediating; Mediation; Mesenchymal Stem Cells; Methods; Modeling; Monitor; Mus; Mutagenesis; Natural regeneration; Nature; Nuclear; Operative Surgical Procedures; Orthopedics; Osteogenesis; Outcome; Pathway interactions; Peptides; 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 therapy; healing; immunogenicity; improved; in vivo; insight; interest; mimetics; nanoGold; nanoparticle; new technology; non-viral gene delivery; novel; osteogenic; plasmid DNA; public health relevance; repaired; scaffold; self assembly; stem cell differentiation; success; tissue repair; trafficking; uptake; viral gene delivery

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）：大段骨缺损是一项持续的临床挑战，会造成巨大的经济成本以及个人健康和社会生产力成本。骨移植的局限性强调需要替代策略。因此，已经探索了无数的治疗方法来解决这一健康问题，包括直接递送生长因子和递送病毒生长因子基因疗法。虽然这些方法改善了愈合结果，但它们尚未在临床上广泛使用，至少部分原因是有限的蛋白质稳定性（直接递送）和安全性问题（病毒基因疗法）。诱导有效的细胞介导生产的非病毒策略 将为克服这些困难提供一个具有挑战性的方法。 我们建议通过开发具有可调DNA结合和可控基因递送的新的组蛋白靶向基因转移支架来解决这些挑战。组蛋白基序在纳米颗粒支架（如纳米金）上的展示将利用我们的初步研究 证明了后修饰的组蛋白尾部促进核积累、DNA释放、转录和增强的非病毒载体转染。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，生物相容性，功能化和细胞进入能力。纳米金上组蛋白尾部的新呈现应该模仿组蛋白八聚体上这些序列的天然、多方面的显示和功能。因此，这些新的支架应该提供额外的重要的，但未开发的，控制和理解基因包装，运输和释放，通过提高利用天然基因转移和运输途径的好处。 因此，该提案的目标是创建和优化多功能“设计者”组蛋白以诱导有效的基因转移，并最终实现用于整形外科和其他应用的改善的组织修复。我们将产生纳米金-质粒DNA（pDNA）组装体，并将确定pDNA结合残基和组蛋白肽的多价呈现是否增强pDNA结合稳定性并改善转染。我们将利用传统的以及纳米金特异性成像分析来阐明非病毒基因转移的关键步骤，并将增强的基因转移效率与生长因子（如骨形态发生蛋白-2（BMP-2））的成骨潜力提高明确联系起来。最后，我们将使用小鼠和大鼠矫形模型来测试BMP-2基因产物增强骨形成和增加缺损修复的速度和功效的能力。我们的方法不仅将阐明机制的细节组蛋白相关基因的贩运，但也将最终将是有用的，适用于骨修复，植入功能化和组织工程的一般生物材料平台。