RANKL and VEGF Signals Mediate Cortical Bone Healing Derived from Structural Autografts: Effective Transfer to Processed Allografts via Immobilized rAAV

• a - DePuy/ Spine; c - LAGeT Inc.

Catastrophic failure of structural allografts used to repair critical defects occurs as a consequence of the dead bone’s inability to remodel and repair microfractures. With a goal of generating a remodeling structural allograft; a murine femoral model utilizing live autografts and processed allografts was developed. Radiographic and histological analyses revealed that autograft heals by endochondral bone formation at the graft-host junction as well as by intramembranous bone formation derived from the graft’s periosteum. Allografts heal only by creeping callus from the host. Gene expression analyses show a significant decrease in RANKL and VEGF expression during allograft healing. Both systemic inhibition with RANK:Fc and anti-VEGF, and also local inhibition with rAAV-OPG and rAAV-sFlt1, demonstrated that RANKL and VEGF are required for autograft healing. To see if the addition of RANKL and VEGF signals could stimulate allograft remodeling and vascularization, we developed a novel gene therapy approach in which recombinant adeno-associated virus (rAAV) can be freeze-dried onto the cortical surface of processed allografts without loosing infectivity. Using allografts containing freeze-dried rAAV-RANKL and rAAV-VEGF we demonstrate that both signals are sufficient to induce allograft remodeling and vascularization, which results in the formation of a new bone collar that spans the entire graft. Addition of either signal alone failed to yield significant effects. In conclusion we find that RANKL and VEGF expression is necessary and sufficient for autograft healing and can be transferred via rAAV to improve healing of structural allografts in a murine model.

• If noted the author indicates something of value received. The codes are identified as: a-research or institutional support; b-miscellaneous funding; c-stock or stock options; d-royalties; e-other financial or material support including consulting. .