Regulated ex vivo regional gene therapy for bone repair using an inducible caspase-9 suicide gene system

Abstract

In order to adapt ex vivo regional gene therapy for clinical applications in orthopaedic surgery, safety issues must be considered. In this study we developed a suicide approach using a dual gene expression two step transcriptional amplification lentiviral vector (LV-TSTA) encoding BMP-2 and an inducible caspase 9 (iC9) system that selectively induces apoptosis upon activation with a chemical inducer of dimerization (CID). Transduction of rat bone marrow stromal cells (RBMSCs) with LV-TSTA-iC9/BMP-2 led to abundant BMP-2 production (90.3 ± 7.9 ng/24 h/106 cells) in vitro and stimulated bone formation in a mouse muscle pouch in the absence of CID. Moreover it was shown that CID could be used to selectively induce apoptosis in iC9-transduced cells both in vitro and in vivo. Double exposure to serial dilutions of CID decreased in vitro production of BMP-2 by 85–87% and Luc activity by 97–99% in iC9/BMP-2 or iC9/Luc-transduced cells respectively. Early administration of CID (Days 0–1 post-op) in mice implanted with iC9/BMP-2-transduced RBMSCs was effective in blocking bone formation, indicating that CID was toxic to the transduced cells. In iC9/Luc-implanted mice, late administration of two doses of CID (Days 27–28 post-op) significantly reduced the luciferase signal. The current study provides proof of concept for the potential clinical application of regulated gene therapy to promote bone repair.