Investigators: Shelley R. Winn, PhD , Department of Surgery, Department of Surgery, Division of Plastic & Reconstructive Surgery

BACKGROUND
Following injury, bone has the ability to regenerate itself to a form and function nearly indistinguishable from the pre-injury state. However, if the injury is beyond a critical limit, recovery will not occur without therapeutic interventions. Autografts and implants with banked bone continue as the treatments of choice, although each exhibits limitations and liabilities. Alternatives have included the utilization of bone-graft substitutes that may incorporate bone derivatives and soluble signaling molecules such as mitogens and morphogens. In addition, an evolving treatment modality, gene therapy, offers an exciting avenue for bone regeneration.

Our laboratories have been developing polymer-engineered gene and cell-based therapies targeting patients with craniofacial osseous malformations. Protein and gene-based delivery of recombinant human bone morphogenetic proteins (BMP's) in preclinical and clinical bone defects have resulted in abundant reports of efficacy, although the megadoses of protein necessary for this response increases the risk of systemic toxicity. BMP-2, -4 and -7 are essential components in bone formation and regeneration. In addition, BMP's can be chemoattractants, and promote differentiation of stem cells into bone forming cells.

We have recently observed osseous healing nearly equivalent to autografts with a polymer-engineered gene therapy composite containing rhBMP-4. High dosing of BMP's can be avoided by using vector DNA containing a BMP gene that is incorporated into reactive cells of the wound environment. Thereafter, the reactive cells will constitutively produce the desired BMP at physiologic levels. Other means to stimulate the in situ activation of genes associated with the process of ossification are under investigation. Future projects will be designed for the optimization of the delivery systems' microarchitecture, vector design and vector DNA release rate kinetics, and this will ultimately enhance the regenerative process and provide an alternative to autografts.

METHODS AND RESOURCES
An appropriate delivery system must be utilized to localize, deliver and protect a preferred biologic. Porous bioresorbable scaffolds mimicking bone architecture are fabricated in our polymer chemistry laboratory. Animal surgeries are performed at OHSU, a fully accredited Animal Care Facility (AALALAC), with supervision provided by Veterinarians certified by the American College of Laboratory Animal Medicine. The histology facility is equipped for soft (paraffin) and hard (methacrylate) tissue processing, immunohistochemistry and in situ hybridization. Portable x-ray equipment is available for radiometric analysis. A dedicated imaging system is utilized for computerized radio- and histomorphometry. Our laboratories also have a dedicated tissue culture facility, a multiplate reader and equipment for assessing gene expression.

ADDITIONAL RESEARCH PROJECTS
The implantation of biodegradable polymers provides a powerful method for delivering high, sustained concentrations of chemotherapeutics for brain and peripheral tumors. Our laboratory has an ongoing collaboration to investigate various formulations to enhance survival in a rodent model of surgically resected glioma. Additional drugs, carrier systems and cancer types are currently under investigation.
Intramyocardial skeletal muscle transplantation (cardiomyoplasty) has been shown experimentally to improve heart function in a number of experimental models of chronically injured myocardium. In addition, a patient with severe ischemic heart failure recently received cardiomyoplasty during a coronary artery bypass grafting procedure. Following this procedure, echocardiography and P.E.T. showed evidence of contraction and viability of the grafted autogenous skeletal muscle cells. Our laboratories are establishing preferred parameters for nominal myoblast yield, purity and survival once transplanted into experimental sites.

PUBLICATIONS
1. Hollinger JO, Winn SR. Growth Factors in Bone Regeneration. Int. J. Oral Maxillofac. Surg., in press.

2. Emerich DF, Winn SR. Immunoisolation Cell Therapy for CNS Diseases. Crit. Rev. Ther. Drug Carrier Sys., in press.

3. Hollinger JO, Winn SR, Sfeir C, Calvert JW, Schmitz JP. Bone Regenerating Options for the Craniofacial Skeleton. In: Craniofacial Surgery, Science & Surgical Technique. K. Lin, R. Ogle, J. Jane (Eds), W.B. Saunders, 2001.

4. Ozaki W, Kawamoto HK. Craniofacial Clefting. In: Craniofacial Surgery, Science & Surgical Technique. K. Lin, R. Ogle, J. Jane (Eds), W.B. Saunders, 2001.

5. Winn SR, Hu Y, Sfeir C, Hollinger JO. Gene Therapy Approaches for Modulating Bone Regeneration. Adv. Drug Deliv. Reviews, 42: 121-138, 2000.

6. Shoichet MS, Winn SR. Cell Delivery to the Central Nervous System. Adv. Drug Deliv. Reviews, 42: 81-102, 2000.

7. Hollinger JO, Winn SR, Bonadio J. Strategies for Tissue Engineering to Address Challenges of the Aging Skeleton. Tissue Engineering, 6(4): 341-350, 2000.

8. Winn SR, Hollinger JO. An Osteogenic Cell Culture System to Evaluate the Cytocompatibility of OsteoSet®, a Calcium Sulfate Bone Void Filler. Biomaterials, 21: 2413-2425, 2000.

9. Emerich DF, Winn SR, Hu Y, Marsh J, Snodgrass P, LaFreniere D, Wiens T, Hassler BP, Bartus RT. Injectable Chemotherapeutic Microspheres and Glioma I: Enhanced Survival Following Implantation into the Cavity Wall of Debulked Tumors. Pharm. Res., 17(7): 767-775, 2000.

10. Emerich DR, Winn SR, Snodgrass P, LaFreniere D, Agostino M, Wiens T, Xiong H, Bartus RT. Injectable Chemotherapeutic Microspheres and Glioma II: Enhanced Survival Following Implantation into Deep Inoperable Tumors. Pharm. Res., 17(7): 776-781, 2000.

11. Ozaki W. Craniosynostosis: Diagnosis and Treatment. Doernbecher Journal, 6(1): 24-29, 2000.

12. Winn SR, Uludag H, Hollinger JO. Carrier Systems for Bone Morphogenetic Proteins. Clin. Orthop. Rel. Res., 367S:S95-S106, 1999.

13. Winn SR, Randolph G, Uludag H, Wong S, Hair GA, Hollinger JO. Establishing an Immortalized Human Osteoprecursor Cell Line: OPC1. J. Bone Min. Res., 14(10): 34-46, 1999.

14. Schmitt JM, Hwang K, Winn SR, Hollinger JO. BMP: An Update on Basic Biology and Clinical Relevance. J. Orthop. Res., 17(2): 269-278, 1999.

15. Winn SR, Schmitt JM, Buck D, Hu Y, Grainger D, Hollinger JO. A Tissue Engineered Bone Biomimetic to Regenerate Calvarial Critical-Sized Defects in Athymic Rats. J. Biomed. Mater. Res., 45(4): 414-421, 1999.