Use of a Stringent Dimerizer-Regulated Gene Expression System for Controlled BMP2 Delivery

Abstract

Gene therapy using constitutively active viral promoters to drive expression of bone morphogenetic proteins (BMPs) has been extensively evaluated as a strategy for inducing bone regeneration. However, this approach offers little control over the concentration, timing, or duration of BMP synthesis. To gain greater control over BMP kinetics, we developed a new inducible system for the controlled expression of BMP2 using a two-component transcription factor that is dimerized with rapamycin (Rap). This approach provided stringent control over BMP2 synthesis with no BMP expression detected in the uninduced state. Rapamycin or the less immunosuppressive analogue, AP21967, rapidly and reversibly induced BMP2 in a dose-dependent manner (range 0.1–10 nM). Subcutaneous implants of fibroblasts containing the Rap-inducible system in syngeneic C57BL/6 mice were highly responsive to ip Rap injection (0.1–1 mg/kg). Peak BMP2 levels were detected within 24 h of a single Rap injection and declined to undetectable levels after 8–10 days. Alternate-day Rap injections (1 mg/kg) for 6 weeks induced subcutaneous ectopic bone formation. Rap-dependent healing of a critical-sized cranial defect was also achieved using this system. This regulated BMP2 expression system will be extremely useful for examining the role of timing and sequence of BMP delivery on bone regeneration.