P01.22GENERATION OF GENETICALLY ENGINEERED INDUCED PLURIPOTENT STEM CELL-DERIVED NEURAL STEM CELLS WITHOUT USING VIRAL VECTORS

Suicide gene therapy using genetically engineered stem cells are is one of the most feasible and promising approaches for glioma therapy. Various stem cells, such as neural and mesenchymal stem cells, have been tested for their tumor tropic activity. We have tested stem cells transduced with the herpes simplex virus-thymidine kinase gene (HSV-TK, suicide gene) encoding a viral thymidine kinase which phosphorylates prodrug ganciclovir (GCV) for experimental glioma in rodents, because the HSV-TK/GCV system generates a potent bystander effect. Recently we found that induced pluripotent stem cells (iPSs) also had tumor tropic activity under both in vitro and in vivo conditions. One of the major limitations of use of HSV-TK gene-trasnduced iPSs in clinical field is the use of virus vectors that randomly integrate into the host genome and are associated with the risk of malignant transformation due to insertional mutagenesis. In the present study, we present a non-viral transfection method for obtaining HSV-TK expressing iPS-derived neural stem cells to circumvent the concerns associated with use of viral vectors. Mouse iPS cells were generated from somatic cells by the plasmid vectors expressing Oct4, Sox2, Klf4, c-Myc and Nanog-GFP-IRES-Puror. We differentiated the mouse iPS cells into neural stem cells and then transfected with the plasmid containing HSV-TK gene using electroporation method. In this way, we obtained HSV-TK gene-trasnduced iPSs-derived neural stem cells without using viral vectors for further pre-clinical expariments of glioma therapy.