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

描述（由申请人提供）：离体局部基因治疗有可能解决与大量骨储备缺乏、大量疤痕组织和有限血管供应相关的困难的骨质流失问题，目前尚无一致满意的解决方案。我们已经证明了这一策略的潜在临床应用，通过使用含有BMP-2 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治愈临界大小的啮齿动物股骨缺损。在最近对Ad-BMP-2与LV-BMP-2治疗骨缺损愈合的比较中，LV-BMP-2治疗获得的骨质量更好。我们假设，较长的BMP表达与慢病毒载体相关，是LV载体获得较好结果的原因，并且这种增加的表达时间可能是在患者具有挑战性的生物环境中实现充分骨修复所必需的。为了将基因疗法应用于临床，它必须具有成本效益，安全，并且必须更好地理解利用转导细胞进行骨修复的生物学原理，以便成功地管理人类骨质流失的各种情况。该提案的目标是评估“同一天”离体方法的临床潜力，这种方法可以很容易地适应于人类使用；了解转导细胞和宿主细胞在骨形成和修复中的作用；使用慢病毒载体评估离体策略的安全性。在Specific Aim 1中提出的研究将比较“当日”体外策略与标准的两步细胞收获和扩增方法的功效。在Specific Aim 2中，在细胞分化特异性启动子的控制下表达GFP的转基因小鼠将被用来描述转导细胞和宿主细胞在临界大小骨缺损愈合中的作用。在特异性目标3中，慢病毒载体体外基因治疗的安全性将通过生物分布试验和器官毒性评估来评估。这些提出的目的应该为骨修复的体外基因治疗的可行性、安全性和生物学提供有价值的信息。这些数据将使我们离实现这种人类使用更近一步。

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. PUBLIC HEALTH RELEVANCE: The ultimate goals of this research proposal are to enhance our knowledge of both the biology of bone repair and regional gene therapy, develop a new cost-effective treatment strategy and assess the safety of this therapy in order to move this technology closer to use in humans. In many cases our present treatment regimens are not adequate to manage difficult bone loss problems. Regional gene therapy has the potential to enhance our ability to treat difficult bone repair problems associated with fracture non-union, revision total joint arthroplasty and spinal fusion.