You are hereOctober 26, 2010 | Pluripotent Stem Cells
Forthcoming Stem Cells Publication – Novel Hyperactive Transposons for Genetic Modification of Induced Pluripotent and Adult Stem Cells: A Non-viral Paradigm for Coaxed Differentiation
By Stuart P Atkinson
Non-viral genetic modification of cells is a relatively inefficient process, be it modifying adult cells or in the generation of induced pluripotent stems cells (iPSC). It has been suggested that transposon-based methods of gene transfer may overcome this limitation, but this technique has been plagued by inefficiency problems. However recent work has utilised in vitro molecular evolution to generate a transposon system derived from the Sleeping Beauty (SB) transposase which shows vastly enhanced transposition capabilities, allowing a significant enhancement of stable gene transfer efficiencies.
This month in Stem Cells, this newly evolved SB transposase (SB100X) is evaluated in a range of clinically relevant cell types; iPSCs, mesenchymal stem cells (MSCs) and muscle stem/progenitor cells. Utilising SB100X resulted in a large improvement in transposition and increased marker gene expression in MSCs and myoblasts, whilst retaining a normal karyotype and differentiation capabilities. Analysis of iPSCs showed similar results; stable expression of the marker transgene observed for over 20 passages with cells remaining pluripotent and maintaining the ability to differentiate across multiple lineages. Further work with iPSCs showed that delivery of the myogenic transcription factor PAX3 into iPSCs drove myogenic differentiation, demonstrating that this method of gene transfer can induce specific differentiation of a pluripotent cell type.
This method of gene transfer is potentially safer than other forms of integrative gene expression due to the lack of gene integration bias and as the activity of the promoter/enhancer of the SB terminal repeat sequences is much lower than that of retrovirus LTRs. However, the authors do note that some undetected deleterious changes could be present, indeed caused by the integrative nature of this system. This suggests that further safety measures and more extensive analysis will be required before systems such as these can be moved into a clinical setting.
Belay E, Mátrai J, Acosta-Sanchez A, et al.
Novel Hyperactive Transposons for Genetic Modification of Induced Pluripotent and Adult Stem Cells: A Non-viral Paradigm for Coaxed Differentiation
Stem Cells Express 2010
Mates L, Chuah MK, Belay E et al.
Molecular evolution of a novel hyperactive Sleeping Beauty transposase enables robust stable gene transfer in vertebrates.
Nat Genet 2009;41:753–761.