Scientific Foundations BMP-9-Transduced Prefabricated Muscular Flaps for the Treatment of Bony Defects Mohamed M. Abdelaal, MD,* Sunil S. Tholpady, PhD,* † Jessica D. Kessler, BS, † Raymond F. Morgan, MD,* Roy C. Ogle, PhD* †‡ Charlottesville, Virginia Autologous bone grafting techniques involve the use of tissues that need to be extracted from healthy sites. This can lead to significant donor site mor- bidity that causes a one-site defect to become a two- site defect. Bone grafts can be especially difficult to manipulate, because bone is a relatively nonmold- able tissue. Furthermore, the inability of a bone graft to contain a transplantable vascular supply also limits the possible size that such a bone graft can be. Because of these limitations, a graft that was moldable with a vascular supply would possess significant advantages in reconstructive applica- tions. In this research, gene therapy techniques were used to create such a graft. An adenovirus expressing BMP-9 was injected into the latissimus dorsi of a nude animal to cause bony differentia- tion of that muscle. Differentiation of the muscle to cartilage in bone was measured by reverse tran- scription polymerase chain reaction and immuno- histochemistry to determine the optimal time of flap elevation. After injection of the BMP-9 virus, the animals were biopsied weekly over a 3-week period. Both bone and cartilage markers were dis- covered in these tissues over the study period. Op- timal flap elevation time was established to be 2 weeks after injection of the virus. Key Words: Bone morphogenetic proteins, bone re- generation, gene therapy T herapeutic options available for large sig- nificant bony defects have evolved through a series of advancements. These phases can best be described as removal, repair, re- placement, and regeneration. In reconstruction of bony defects, the current standard of treatment is replacement using autologous bone or bone substi- tutes. These methods suffer from well-known draw- backs, however, including resorption, infection, and second-site morbidity. Regenerative options are con- tinually being expanded, and permutations of gene therapy, matrix biology, and cellular biology will eventually lead to optimal solutions. From a cell biology/soluble factor perspective, some of the most important molecules in bone for- mation and maintenance are the bone morphoge- netic proteins (BMPs). The activity of the BMPs was first described when fractions of demineralized bone matrix were injected into ectopic sites and subse- quently induced bone formation. 1–3 The purified fac- tors most important for this activity were determined to be the BMPs. These cytokines are members of the transforming growth factor- class of molecules and make significant contributions not only during bone healing but also in the formation and determination of several embryological structures. 4–7 Although the BMPs provide a powerful method by which to stimulate the ectopic formation of bone, repeated applications are required to produce the de- sired outcome. One possible solution is the modifi- cation of cells such that they produce native BMPs. This approach has several advantages depending on the method used to alter cells. Insertion of genes into cells is more economical than the use of recombinant protein. Protein production can be modulated in cells to ideal levels and is continuous, whereas bolus in- jections cause fluctuations in the quantity of active cytokine. In this study, a combination of replacement and regeneration is described. A muscle flap from the latissimus dorsi was first injected with an adenovirus expressing BMP-9. Bolus injections were not used because of the inherent limitations of recombinant From the Departments of *Plastic and Reconstructive Surgery, † Cell Biology, and ‡ Neurosurgery, University of Virginia Health Sciences Center, Charlottesville, Virginia. This study was supported by a grant from the National Insti- tutes of Dental and Craniofacial Research (NIDCR), National In- stitutes of Health, R21 DE 15023-02. Address correspondence and reprint requests to Dr Ogle, De- partment of Plastic and Reconstructive Surgery, University of Vir- ginia Health Sciences Center, Charlottesville, VA 22903. E-mail: rco2j@virginia.edu 736