High expression of human clotting factor IX cDNA in myoblasts C2C12 cells and C3H mice

Mouse myoblast C2C12 cell was used as target cell for gene transfer study of human clotting factor IX (hFIX) cDNA. In addition to the previously constructed retroviral vectors XLIX, LNCIX and G1NaCIX, G1NaMCIX with hFIX driven by muscle creatine kinase (MCK) enhancer and human cytomegalovirus (CMV) was constructed, based on the retroviral vector G1Na. These four retroviral vectors were used to transduce mouse myoblasts C2C12. With ELISA assays, it has been found that the expression levels of human clotting factor IX detected in those transduced C2C12 cells are G1NaMCIX> G1NaCIX> LNCIX> XLIX. Mixed colonal cells transduced with GlNaMCIX expressed hFIX protein at the level of 640 ng/10(6) cell every 24 h. The modified C2C12 cells transduced with G1NaMCIX were implanted into skeletal muscle of the hindlegs of C3H mice; a stable expression of hFIX was detected and lasted for 35 d, with a maximum level of 206 ng/mL plasma. The regulation of hFIX cDNA expression in myoblasts was discussed and it was strongly suggested that a myoblast-mediated gene delivery system had the potential to be optimized as a safe and effective therapeutic modality for hemophilia B.