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Transposition from a gutless adeno-transposon vector stabilizes transgene expression in vivo

Nature Biotechnology volume 20, pages 999–1005 (2002)Cite this article

Abstract

A major limitation of adenovirus-mediated gene therapy for inherited diseases is the instability of transgene expression in vivo, which originates at least in part from the loss of the linear, extrachromosomal vector genomes. Herein we describe the production of a gene-deleted adenovirus–transposon vector that stably maintains virus-encoded transgenes in vivo through integration into host cell chromosomes. This system utilizes a donor transposon vector that undergoes Flp-mediated recombination and excision of its therapeutic payload in the presence of the Flp and Sleeping Beauty recombinases. Systemic in vivo delivery of this system resulted in efficient generation of transposon circles and stable transposase-mediated integration in mouse liver. Somatic integration was sufficient to maintain therapeutic levels of human coagulation Factor IX for more than six months in mice undergoing extensive liver proliferation. These vectors combine the versatility of adenoviral vectors with the integration capabilities of a eukaryotic DNA transposon and should prove useful in the treatment of genetic diseases.

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Figure 1: Analysis of transposition from E1/E3-deleted adenoviruses and linear transposon DNA.
Figure 2: The adeno-transposon system.
Figure 3: Conditional rearrangement and chromosomal integration of HD vectors in vivo following expression of the Flp and SB recombinases.
Figure 4: Adenovirus-based β-galactosidase expression in mouse liver before and after hepatocellular regeneration.
Figure 5: In vivo human Factor IX persistence in actively dividing mouse livers via the adeno-transposon system.
Figure 6: Transposition in vivo from a single Ad vector encoding transposon and Flp-activated transposase activities.

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Acknowledgements

We thank K. Ohashi for helpful discussions, J. Chamberlain for providing the C7-Cre cells, and S. Dymecki for providing Flp/FRT-based plasmids. This work was supported by NIH grant DK49022 (M.A.K.). A.E. is the recipient of a Judith Pool National Hemophilia Fellowship. J.G.M. was funded by grants from the Carlsberg Foundation and the Danish Medical Research Council.

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  1. Department of Pediatrics and Genetics, Stanford University School of Medicine, Stanford, 94305-5208, CA

    Stephen R. Yant, Anja Ehrhardt, Jacob Giehm Mikkelsen, Leonard Meuse, Thao Pham & Mark A. Kay

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Yant, S., Ehrhardt, A., Mikkelsen, J. et al. Transposition from a gutless adeno-transposon vector stabilizes transgene expression in vivo. Nat Biotechnol 20, 999–1005 (2002). https://doi.org/10.1038/nbt738

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