A paper summarizing 18 years of laboratory work and research to develop a “same day” regional gene therapy to enhance bone repair was the recipient of the Kappa Delta Elizabeth Winston Lanier Award. The paper, “Regional Gene Therapy (RGT) to Enhance Bone Repair,” by Jay R. Lieberman, MD, describes progress in research into this type of therapy as part of a comprehensive tissue engineering strategy to treat patients with large bone defects or a compromised biologic environment.

According to Dr. Lieberman, the method could be used for treatment in situations such as fracture nonunion, spinal fusion, and revision total joint arthroplasty. Current procedures to treat large bone defects or enhance bone repair in a compromised biologic environment are associated with morbidity and high costs.

Encouraging bone repair
Four elements are critical for bone repair, noted Dr. Lieberman: osteoinductive signals, the availability of osteogenic cells capable of responding to these signals, an osteoconductive matrix, and an adequate vascular supply. Currently, recombinant bone morphogenetic proteins (rhBMPs) are the only osteoinductive agents available; they have been approved by the U.S. Food and Drug Administration for use in fresh tibial fractures and spinal fusion.

BMPs induce healing by promoting mesenchymal cell proliferation and osteoprogenitor differentiation into osteoblasts. Dr. Lieberman notes, however, “in humans, supraphysiologic doses of the protein are required to produce adequate bone formation. Thus, milligram doses of rhBMP are needed to induce bone repair, but there are only nanogram amounts of these proteins in the human body.”

This suggests that BMPs are being used in an inefficient fashion at the implantation site. High doses are expensive and may contribute to some of the side effects that have been associated with BMP use, including ectopic bone formation and soft-tissue edema.

Autogenous bone graft can be effective, but the supply is limited and graft harvest has associated morbidities, including pain, risk of infection, and nerve injury.

Regional gene therapy
Regional gene therapy is a novel approach that incorporates the transfer of genetic information for therapeutic purposes, functioning as a “biological protein delivery system.”

“We have tried to develop a regimen that will meet the following criteria: enhance the biology of bone repair, facilitate ease of use by the surgeon, promote safety, and be cost-effective,” said Dr. Lieberman.

His laboratory was the first to heal critical-sized defects in a rat using an ex vivo strategy, with BMP-2–producing bone marrow cells created via adenoviral or lentiviral gene transfer. However, he noted, the cell expansion step involved in traditional ex vivo therapy makes it expensive and time-consuming.

In traditional ex vivo gene therapy, host cells are harvested from a specific anatomic state and expanded in tissue culture. DNA sequences are then transferred into the cells in tissue culture and the cells are reimplanted at the desired location. With in vivo therapy, transfer occurs within the host with direct administration of the vector—viral (transduction) or nonviral (transfection)—systemically or locally at the specific anatomic site.

In the “same day” regimen, cell harvest, cell transduction, and implantation occur at the same sitting. “It has been demonstrated that the ‘same day’ gene therapy produced better bone quality on micro CT and biomechanical testing when compared to the traditional two-step ex vivo approach,” Dr. Lieberman said.

Although the in vivo strategy was attractive due to its simplicity, concerns about its efficacy were raised. Because RGT would be used in difficult bone repair scenarios, the number of responding cells at the repair site might be insufficient to transduce and produce the appropriate osteoinductive response.

The ex vivo strategy had several potential advantages, including the following:

• The surgeon’s ability to select a specific cellular delivery vehicle (ie, stem cells, bone marrow cells, muscle cells, skin fibroblasts) for a particular clinical scenario
• The available choice of different carriers for the transduced cells, depending on the anatomic location of the defect and the biomechanic requirements necessary to create a stable bone construct
• Possibly better safety than with an in vivo approach with viral vectors because no viral complexes of DNA are injected directly into the body. However, further study of the lentiviral vector is necessary.

Dr. Lieberman and his colleagues decided to use lentiviral vectors because of the prolonged protein production, their ability to infect dividing and nondividing cells, and their minimal immunogenicity. Although lentiviral vectors have their origin in human immunodeficiency virus (HIV), which was a safety concern, better bone repair was noted with lentiviral than adenoviral gene transfer.

In working on the lentiviral technique, the researchers modified the standard vector using a two-step transcriptional amplification system, which was found to produce significantly more BMP after cell transduction than the standard lentiviral vector.

In rats, same-day RGT was successful in bone repair, with biomechanic testing results equivalent to those of unoperated control femurs. “We are excited by our results with the ‘same-day’ strategy because this regimen is convenient and may be cost-effective,” said Dr. Lieberman. “Our goal now is to continue to adapt “same-day” regional gene therapy for clinical use in patients.”

Disclosures: Dr. Lieberman—DePuy, A Johnson & Johnson Company; Amgen; Arthrex; Journal of Arthroplasty, Journal of the AAOS, American Association of Hip and Knee Surgeons.

Terry Stanton is the senior science writer for AAOS Now; he can be reached at tstanton@aaos.org

Bottom Line

• Bone morphogenetic proteins are potent osteoinductive agents, but are costly and have associated side effects.
• Autologous bone graft can be effective, but the supply is limited, and morbidity is associated with graft harvest.
• Previous gene therapy techniques involved time and expense of expansion.
• In “same-day” regional gene therapy (RGT), all the steps of culture and gene delivery are done in one setting.
• Successful results in rats hold promise for clinical use of same-day RGT.