The Use of Regional Gene Delivery to Heal Critical Sized Bone Defects

描述（由申请人提供）：离体区域基因治疗具有解决与大骨量缺乏、大量瘢痕组织和有限血管供应相关的困难的骨丢失问题的潜力，目前还没有一致满意的解决方案。我们已经通过用含有BMP-2的cDNA的腺病毒载体（Ad-BMP-2）或慢病毒载体（LV-BMP-2）转导的大鼠和人骨细胞成功愈合临界尺寸的啮齿动物股骨缺损，证明了这种策略的潜在临床实用性的概念证明。在最近用Ad-BMP-2与LV-BMP-2治疗骨缺损愈合的比较中，用LV-BMP-2治疗获得了更好质量的骨。我们假设，与慢病毒载体相关的BMP表达时间更长是LV载体获得更好结果的原因，并且这种表达持续时间的增加可能是在患者中观察到的挑战性生物环境中实现充分骨修复所必需的。为了使基因疗法被用于临床应用，它必须具有成本效益，安全，并且必须更好地理解用转导细胞进行骨修复的生物学，以便成功地管理在人类中观察到的广泛变异性骨丢失情况。该提案的目标是评估“同一天”离体方法的临床潜力，该方法可以很容易地适用于人类使用;了解转导细胞和宿主细胞在骨形成和修复中的作用;评估使用慢病毒载体的离体策略的安全性。具体目标1中提出的研究将比较“同一天”ev vivo策略与标准两步细胞收获和扩增方法的功效。在特异性目的2中，在细胞分化特异性启动子的控制下表达GFP的转基因小鼠将用于描述转导细胞和宿主细胞在关键尺寸骨缺损愈合中的作用。在特定目标3中，将通过生物分布测定和器官毒性评价来评估使用慢病毒载体的离体基因治疗的安全性。这些建议的目的应该提供有关的可行性，安全性和生物学的离体基因治疗骨修复的有价值的信息。这些数据应该使我们更接近实现这种人类用途。

DESCRIPTION (provided by applicant): Ex vivo regional gene therapy has the potential to solve difficult bone loss problems associated with large bone stock deficiency, abundant scar tissue and limited vascular supply for which at the present time there is no consistently satisfactory solution. We have demonstrated proof of concept the potential clinical utility of this strategey by successfully healing critical sized rodent femoral defects with rat and human bone cells transduced with either an adenoviral vector (Ad- BMP-2) or lentiviral vector (LV-BMP-2) containing the cDNA for BMP-2. In a recent comparison of healing of a bone defect with Ad-BMP-2 versus LV-BMP-2 better quality bone was obtained with the LV-BMP-2 treatment. We hypothesized that the more prolonged BMP expression associated with the lentiviral vector was responsible for the better results with the LV vector and that this increased duration of expression may be necessary to achieve adequate bone repair in the challenging biological enviorments seen in patients. In order for gene therapy to be adopted for clinical use it must be cost-effective, safe and there must be a better understanding of the biology of bone repair with transduced cells in order to successfully manage the wide variability bone loss scenarios seen in humans. The goals for this proposal are to assess the clinical potential of a "same day" ex vivo approach that can be easily adapted for human use; understand the role of transduced cells and host cells in bone formation and repair and; evaluate the safety of an ex vivo strategy using a lentiviral vector. The research proposed in Specific Aim 1 will compare the efficacy of a "same day" ev vivo strategy with the standard two step cell harvest and expansion approach. In Specific Aim 2 transgenic mice that express GFP under the control of cell differentiation specific promoters will be used to delineate the role of transduced cells and host cells in the healing of criticial sized bone defects. In Specific Aim 3 the safety of ex vivo gene therapy with a lentiviral vector will be assessed with a biodistribution assay and an evaluation of organ toxicity. These proposed Aims should provide valuable information regarding the feasibility, saftey, and biology of ex vivo gene therapy for bone repair. This data should move us one step closer to implementing this human use.