The application of genetic engineering techniques for the induction of bone formation currently being evaluated in pre-clinical and clinical studies for the treatment of numerous pathologic disorders, including osteoporosis and nonhealing long bone fractures. Bone repair and reconstruction also has significant implications for improving the healing of facial fractures, the restoration of bony tissues in developmental craniofacial disorders, and the regeneration of alveolar bone in periodontal disease. The induction of osteogenesis throughout the skeletal system using bone morphogenetic proteins (BMPs), typically on a biologic carrier, has had success in the experimental setting, and is now being tested for clinical use. Our laboratory has recently demonstrated that direct BMP gene transfer techniques can also be utilized to stimulate bone formation in ectopic and orthotopic regions, including critical size mandibular defects. However, first generation BMP adenoviral vectors currently in use are limited by the immune responses they elicit in immunocompetent animals. The engineering of novel BMP viral vectors, however, should advance the clinical efficacy of this unique technique for the treatment disorders involving bone loss. We hypothesize in this grant proposal that the efficacy of BMP genetic therapy can be enhanced by: 1) the generation of novel viral vectors which improve transgene expression, 2) the use of tissue-specific control elements to localize BMP expression to specific target cells, 3) the use of an inducible expression system to temporally regulate BMP production within target cells, and 4) the utilization of ex vivo gene therapy techniques. To test these hypotheses, we propose the following studies: I. To decrease the antigenicity of the first generation adenoviral vectors and increase the length of transgene expression, we will construct, produce, and test two novel BMP vectors, including a gutless adenoviral vector and an adeno-associated viral vector. II. To restrict BMP expression to specific cell populations, we will produce and test BMP viral vectors with the transgene under control of the osteocalcin promoter (osteoblast specific). III. To externally regulate osteogenesis, we will produce and test a BMP viral vector with the transgene under control of tetracycline responsive control elements. IV. To evaluate ex vivo gene therapy, human mesenchymal stem cells will be transduced with BMP genes and delivered on a Type I collagen scaffold. The osteogenic activity of these novel approaches will be evaluated in a rigorous fashion using immunohistochemistry, histology, and computed tomography in heterotopic and orthotopic locations, providing the scientific foundation for future basic and clinical BMP gene therapy studies.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR046488-02
Application #
6621538
Study Section
Special Emphasis Panel (ZRG1-SSS-M (01))
Program Officer
Panagis, James S
Project Start
2002-03-01
Project End
2004-11-30
Budget Start
2003-03-01
Budget End
2003-11-30
Support Year
2
Fiscal Year
2003
Total Cost
$235,690
Indirect Cost
Name
University of Virginia
Department
Neurosurgery
Type
Schools of Medicine
DUNS #
065391526
City
Charlottesville
State
VA
Country
United States
Zip Code
22904
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Li, J Z; Li, H; Hankins, G R et al. (2005) Local immunomodulation with CD4 and CD8 antibodies, but not cyclosporine A, improves osteogenesis induced by ADhBMP9 gene therapy. Gene Ther 12:1235-41
Li, Jin Zhong; Holman, David; Li, Hongwei et al. (2005) Long-term tracing of adenoviral expression in rat and rabbit using luciferase imaging. J Gene Med 7:792-802
Li, Jin Zhong; Li, Hongwei; Dunford, Brad et al. (2003) Rat strain differences in the ectopic osteogenic potential of recombinant human BMP adenoviruses. Mol Ther 8:822-9
Li, J Z; Li, H; Sasaki, T et al. (2003) Osteogenic potential of five different recombinant human bone morphogenetic protein adenoviral vectors in the rat. Gene Ther 10:1735-43
Li, Jin Zhong; Hankins, Gerald R; Kao, Chinghai et al. (2003) Osteogenesis in rats induced by a novel recombinant helper-dependent bone morphogenetic protein-9 (BMP-9) adenovirus. J Gene Med 5:748-56
Dayoub, Hayan; Dumont, Randall J; Li, Jin Zhong et al. (2003) Human mesenchymal stem cells transduced with recombinant bone morphogenetic protein-9 adenovirus promote osteogenesis in rodents. Tissue Eng 9:347-56
Jane Jr, John A; Dunford, Bradley A; Kron, Adam et al. (2002) Ectopic osteogenesis using adenoviral bone morphogenetic protein (BMP)-4 and BMP-6 gene transfer. Mol Ther 6:464-70
Dumont, Randall J; Dayoub, Hayan; Li, Jin Zhong et al. (2002) Ex vivo bone morphogenetic protein-9 gene therapy using human mesenchymal stem cells induces spinal fusion in rodents. Neurosurgery 51:1239-44; discussion 1244-5