Baculovirus-Mediated Gene Delivery for Bone Engineering

Abstract

Baculovirus has emerged to be a promising gene delivery vector and adipose-derived stem cells have captured interests for bone regeneration. Massive segmental defects arising from trauma or tumor resection remain a challenging clinical problem. To heal massive, segmental bone defects using adipose derived stem cells (ASCs), which alone cannot heal large defects, we hypothesized that sustained expression of factors promoting bone regeneration (BMP2) and angiogenesis (VEGF) provides continuous stimuli to augment the healing. Insect baculovirus (BV) holds promise for gene therapy and efficiently transduces stem cells, but it only mediates transient transgene expression. Therefore we developed a dual BV system whereby one BV expressed FLP recombinase (BacFLP) while the other hybrid BV harbored an Frt-flanking transgene cassette. After sequential transduction with BacFLP and the hybrid BV, FLP/Frt-mediated recombination occurred in up to 46% of ASCs, leading to cassette excision off the BV genome, formation and persistence of episomal transgene and prolongation of expression to >;28 days. Transduction of ASCs with the BMP2-encoding hybrid BV led to prolonged BMP2 expression and augmented ASCs osteogenesis even without other osteogenic supplements. ASCs engineered by the hybrid vectors mediating sustained BMP2/VEGF expression healed the critical-size (10 mm) segmental bone defects in 12 out of 12 New Zealand White rabbits in 8 weeks, which significantly outperformed ASCs engineered with BV transiently expressing BMP2/VEGF with respect to healing rate, bone metabolism, bone volume, bone density, angiogenesis and mechanical properties. These data confirmed our hypothesis that persistent BMP2/VEGF expression is essential. The use of the hybrid BV vector represents a novel therapy to treat massive segmental defects necessitating concerted ossification and vascularization.