Healing of Segmental Defects of Bone by Gene Transfer

基因转移修复节段性骨缺损

基本信息

批准号：
8693923
负责人：
CHRISTOPHER Howard EVANS
金额：
$ 61.74万
依托单位：
MAYO CLINIC ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2003
资助国家：
美国
起止时间：
2003-08-01 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8693923
关键词：
Actins Address Adenovirus Vector Adenoviruses Animal Model Animals Attenuated Autologous Autopsy Biodistribution Biological Biology Blood Vessels Calcified Cartilage Cells Chemotaxis Chondrocytes Clinical Collaborations Complementary DNA Contralateral DNA Data Defect Deposition Development Diaphyses Dose Effectiveness Endothelial Cells Euthanasia Evaluation Fatty acid glycerol esters Feasibility Studies Femur Fluoroscopy Fracture Funding Gene Expression Gene Transfer Genes Genetic Genetic Enhancement Genetically Modified Animals Goals Grant Harvest Healed Histology Histopathology Human Immune Immune response Immune system Immunosuppressive Agents Implant In Vitro Inbred F344 Rats Infection Inferior Injection of therapeutic agent LacZ Genes Lesion Luc Gene Luciferases Mechanics Mediating Methods Modality Modeling Morphology Mus Muscle Muscle Cells Myosin ATPase Nude Rats Operative Surgical Procedures Organ Osteoblasts Osteocalcin Osteogenesis Osteotomy Population Positioning Attribute Procedures Process Production Progress Reports Property Radiology Specialty Rattus Reaction Recombinants Reporter Reporter Genes Research Research Institute Role Rosa Series Sheep Skeletal Muscle Staining method Stains Stem cells Steroids Switzerland Technology Testing Thick Time TimeLine Transcriptional Regulation Transgenes Translational Research Vascular Endothelial Cell Viral Genome Virus Work X-Ray Computed Tomography Xenograft procedure base bone bone cell bone healing bone morphogenetic protein 2 cell type gene therapy head-to-head comparison healing implantation improved in vivo in vivo imaging insight long bone muscle form new technology novel offspring osteogenic osteoprogenitor cell programs promoter recombinant human bone morphogenetic protein-2 recombinase research study response safety testing scaffold tissue repair transgene expression vector

项目摘要

DESCRIPTION (provided by applicant): This competing renewal seeks to continue our development of novel, gene-based methods for improving the healing of large segmental defects in bone. For reasons discussed in the body of the grant, a recombinant adenovirus vector carrying the cDNA for human bone morphogenetic protein-2 (Ad.BMP-2) is used for this project. During the first funding cycle of this grant, using a rat femoral defect model, we noted a remarkable improvement in defect healing when skeletal muscle from syngeneic rats was transduced with Ad.BMP-2 and then inserted into the defect. Under these conditions, healing of the defect was rapid, reliable and uniform. In the next cycle of this grant we propose two Specific Aims that will: i) establish the biology that underlies this phenomenon and ii) evaluate the efficacy of the technology in a large animal model, the sheep. The latter Aim will be completed in collaboration with AO Research Institute in Davos, Switzerland. Specific Aim 1 will test the hypothesis that the high effectiveness of Ad.BMP-2 modified muscle grafts in promoting bone healing reflects the ability of the graft to supply not only endogenously synthesized BMP-2, but also progenitor cells that form cartilage and then bone. Because immune responses to adenovirus is a recognized problem for certain applications of adenovirus-based gene therapy, Fischer rats will be used to determine the duration of transgene expression in the rat and the role of the immune system in curtailing expression. Based upon our preliminary data, the abbreviated ex vivo method we have developed eliminates the humoral response to the virus. To enable us to take advantage of genetically modified mice to address the other questions raised in Specific Aim 1, we have developed a new model in which muscle from genetically modified mice are inserted into femoral defects in athymic rats. Using this model we will insert Ad.BMP-2 transduced muscle grafts from the following genetically modified mice: mice containing the luciferase gene under the control of the human osteocalcin promoter; Rosa mice, all of whose cells are LacZ+; and mice derived from crosses between Tie2- Cre mice and R26R mice to enable lineage analysis of tie2+ cells derived from endothelial cells of blood vessels. The sheep studies will utilize a standard 3cm, critical size tibial defect to evaluate the reparative ability of Ad.BMP-2 transduced autologous muscle grafts. Healing will be assessed by serial radiology. After 6 months, sheep will be euthanized and healing assessed by micro-computed tomography, histology, mechanical testing and intravital fluoroscopy. The immune response of the sheep to the vector will also be assessed, and various organs will be recovered at necropsy for evaluation of histopathology and the distribution of viral genomes.

描述（由申请人提供）：本次竞争更新旨在继续开发基于基因的新颖方法，以改善骨中大节段缺损的愈合。由于资助正文中讨论的原因，该项目使用了携带人骨形态发生蛋白 2 (Ad.BMP-2) cDNA 的重组腺病毒载体。在本次资助的第一个资助周期中，我们使用大鼠股骨缺损模型，注意到当用 Ad.BMP-2 转导同系大鼠的骨骼肌然后将其插入缺损处时，缺损愈合有显着改善。在这些条件下，缺损的愈合快速、可靠且均匀。在这笔拨款的下一个周期中，我们提出了两个具体目标：i）建立这一现象背后的生物学原理；ii）评估该技术在大型动物模型（绵羊）中的功效。后一个目标将与瑞士达沃斯 AO 研究所合作完成。具体目标 1 将检验这样的假设：Ad.BMP-2 修饰的肌肉移植物在促进骨愈合方面的高效性反映了移植物不仅提供内源合成的 BMP-2，而且还提供形成软骨和骨的祖细胞的能力。由于对腺病毒的免疫反应是基于腺病毒的基因治疗的某些应用中公认的问题，因此费舍尔大鼠将用于确定大鼠中转基因表达的持续时间以及免疫系统在抑制表达中的作用。根据我们的初步数据，我们开发的简化离体方法消除了对病毒的体液反应。为了使我们能够利用转基因小鼠来解决特定目标 1 中提出的其他问题，我们开发了一种新模型，将转基因小鼠的肌肉插入无胸腺大鼠的股骨缺损中。使用该模型，我们将插入来自以下转基因小鼠的 Ad.BMP-2 转导的肌肉移植物：含有受人骨钙素启动子控制的荧光素酶基因的小鼠； Rosa小鼠，所有细胞均为LacZ+； Tie2-Cre 小鼠和 R26R 小鼠杂交衍生的小鼠，可对源自血管内皮细胞的 ti2+ 细胞进行谱系分析。绵羊研究将利用标准的 3 厘米临界尺寸胫骨缺损来评估 Ad.BMP-2 转导的自体肌肉移植物的修复能力。将通过连续放射学评估愈合情况。 6 个月后，将对羊实施安乐死，并通过微型计算机断层扫描、组织学、机械测试和活体荧光检查评估愈合情况。还将评估绵羊对载体的免疫反应，并在尸检时回收各种器官，以评估组织病理学和病毒基因组的分布。

项目成果

期刊论文数量（25）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Extended release of proteins following encapsulation in hydroxyapatite/chitosan composite scaffolds for bone tissue engineering applications.

DOI：
10.1016/j.msec.2017.11.001
发表时间：
2018-03-01
期刊：
Materials science & engineering. C, Materials for biological applications
影响因子：
0
作者：
Devine DM;Hoctor E;Hayes JS;Sheehan E;Evans CH
通讯作者：
Evans CH

Altered relative expression of BMPs and BMP inhibitors in cartilaginous areas of human fractures progressing towards nonunion.

DOI：
10.1002/jor.20794
发表时间：
2009-06
期刊：
JOURNAL OF ORTHOPAEDIC RESEARCH
影响因子：
2.8
作者：
Kwong, Francois N. K.;Hoyland, Judith A.;Freemont, Anthony J.;Evans, Christopher H.
通讯作者：
Evans, Christopher H.

Application of structural rigidity analysis to assess fidelity of healed fractures in rat femurs with critical defects.

DOI：
10.1007/s00223-010-9353-4
发表时间：
2010-05
期刊：
CALCIFIED TISSUE INTERNATIONAL
影响因子：
4.2
作者：
Nazarian, Ara;Pezzella, Lina;Tseng, Alan;Baldassarri, Stephen;Zurakowski, David;Evans, Christopher H.;Snyder, Brian D.
通讯作者：
Snyder, Brian D.

Advances in regenerative orthopedics.

DOI：
10.1016/j.mayocp.2013.04.027
发表时间：
2013-11
期刊：
Mayo Clinic proceedings
影响因子：
8.9
作者：
Evans CH
通讯作者：
Evans CH

Evaluation of BMP-2 gene-activated muscle grafts for cranial defect repair.

DOI：
10.1002/jor.22038
发表时间：
2012-07
期刊：
JOURNAL OF ORTHOPAEDIC RESEARCH
影响因子：
2.8
作者：
Liu, Fangjun;Porter, Ryan M.;Wells, James;Glatt, Vaida;Pilapil, Carmencita;Evans, Christopher H.
通讯作者：
Evans, Christopher H.