Design of a nonviral vector for site-selective, efficient integration into the human genome.

Gene therapy in eukaryotes has met many obstacles. Research into the design of suitable nonviral vectors has been slow. To our knowledge, no nonviral vector has been proposed that allows for the possibility of highly efficient, site-selective integration into the genome of mammalian cells. On the basis of prior studies investigating the components necessary for transposon, retrovirus-like retrotransposon, and retroviral integration, we propose a nonviral system that would potentially allow for site-selective, efficient integration into the mammalian genome. Transposons have been developed that can transform a variety of cell lines. For example, the Sleeping Beauty transposon (SB) can transform a wide range of vertebrate cells from fish to human, and it mediates stable integration and long-term transgene expression in mice. However, the efficiency of transposition varies significantly among cell lines, suggesting the possible involvement of host factors in SB transposition. Here, we propose the use of a chimeric transposase (i.e., transposase-host DNA binding domain) to bypass the potential requirement of a host DNA-directing factor (or factors) for efficient, site-selective integration. We also discuss another potential method of docking the transposon-based vector adjacent to the host DNA, utilizing repetitive sequences for homologous recombination to promote efficient site-selective integration, as well as other site-selective nonviral approaches.