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Extending the transposable payload limit of Sleeping Beauty (SB) using the Herpes Simplex Virus (HSV)/SB amplicon-vector platform

Gene Therapy volume 17, pages 424–431 (2010)Cite this article

Abstract

The ability of a viral vector to safely deliver and stably integrate large transgene units (transgenons), which not only include one or several therapeutic genes, but also requisite native transcriptional regulatory elements, would be of significant benefit for diseases presently refractory to available technologies. The herpes simplex virus type-1 (HSV-1) amplicon vector has the largest known payload capacity of approximately 130 kb, but its episomal maintenance within the transduced cell nucleus and induction of host cell silencing mechanisms limits the duration of the delivered therapeutic gene(s). Our laboratory developed an integration-competent version of the HSV-1 amplicon by adaptation of the Sleeping Beauty (SB) transposon system, which significantly extends transgene expression in vivo. The maximum size limit of the amplicon-vectored transposable element remains unknown, but previously published plasmid-centric studies have established that DNA segments longer than 6-kb are inefficiently transposed. Here, we compared the transposition efficiency of SB transposase in the context of both the HSV amplicon vector as well as the HSV amplicon plasmid harboring 7 and 12-kb transposable reporter transgene units. Our results indicate that the transposition efficiency of the 12-kb transposable unit via SB transposase was significantly reduced as compared with the 7-kb transposable unit when the plasmid version of the HSV amplicon was used. However, the packaged HSV amplicon vector form provided a more amenable platform from which the 12-kb transposable unit was mobilized at efficiency similar to that of the 7-kb transposable unit via the SB transposase. Overall, our results indicate that SB is competent in stably integrating transgenon units of at least 12 kb in size within the human genome upon delivery of the platform via HSV amplicons.

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References

  1. Hermonat PL, Quirk JG, Bishop BM, Han L . The packaging capacity of adeno-associated virus (AAV) and the potential for wild-type-plus AAV gene therapy vectors. FEBS Lett 1997; 407: 78–84.

    Article  CAS  Google Scholar 

  2. Kumar M, Keller B, Makalou N, Sutton RE . Systematic determination of the packaging limit of lentiviral vectors. Hum Gene Ther 2001; 12: 1893–1905.

    Article  CAS  Google Scholar 

  3. Senior SL, Wade-Martins R . Herpes simplex virus type 1 amplicon vectors for the infectious delivery and expression of genomic DNA loci. Curr Opin Mol Ther 2005; 7: 337–345.

    CAS  PubMed  Google Scholar 

  4. Suzuki M, Kasai K, Saeki Y . Plasmid DNA sequences present in conventional herpes simplex virus amplicon vectors cause rapid transgene silencing by forming inactive chromatin. J Virol 2006; 80: 3293–3300.

    Article  CAS  Google Scholar 

  5. Wang Y, Camp SM, Niwano M, Shen X, Bakowska JC, Breakefield XO et al. Herpes simplex virus type 1/adeno-associated virus rep(+) hybrid amplicon vector improves the stability of transgene expression in human cells by site-specific integration. J Virol 2002; 76: 7150–7162.

    Article  CAS  Google Scholar 

  6. Heister T, Heid I, Ackermann M, Fraefel C . Herpes simplex virus type 1/adeno-associated virus hybrid vectors mediate site-specific integration at the adeno-associated virus preintegration site, AAVS1, on human chromosome 19. J Virol 2002; 76: 7163–7173.

    Article  CAS  Google Scholar 

  7. Ivics Z, Hackett PB, Plasterk RH, Izsvak Z . Molecular reconstruction of Sleeping Beauty, a Tc1-like transposon from fish, and its transposition in human cells. Cell 1997; 91: 501–510.

    Article  CAS  Google Scholar 

  8. Bowers WJ et al. Neuronal precursor-restricted transduction via in utero CNS gene delivery of a novel bipartite HSV amplicon/transposase hybrid vector. Mol Ther 2006; 13: 580–588.

    Article  CAS  Google Scholar 

  9. Peterson EB, Mastrangelo MA, Federoff HJ, Bowers WJ . Neuronal specificity of HSV/sleeping beauty amplicon transduction in utero is driven primarily by tropism and cell type composition. Mol Ther 2007; 15: 1848–1855.

    Article  CAS  Google Scholar 

  10. Izsvak Z, Ivics Z, Plasterk RH . Sleeping Beauty, a wide host-range transposon vector for genetic transformation in vertebrates. J Mol Biol 2000; 302: 93–102.

    Article  CAS  Google Scholar 

  11. Karsi A, Moav B, Hackett P, Liu Z . Effects of insert size on transposition efficiency of the sleeping beauty transposon in mouse cells. Mar Biotechnol (NY) 2001; 3: 241–245.

    Article  CAS  Google Scholar 

  12. Geurts AM, Yang Y, Clark KJ, Liu G, Cui Z, Dupuy AJ et al. Gene transfer into genomes of human cells by the sleeping beauty transposon system. Mol Ther 2003; 8: 108–117.

    Article  CAS  Google Scholar 

  13. Zayed H, Izsvak Z, Walisko O, Ivics Z . Development of hyperactive sleeping beauty transposon vectors by mutational analysis. Mol Ther 2004; 9: 292–304.

    Article  CAS  Google Scholar 

  14. Yant SR, Huang Y, Akache B, Kay MA . Site-directed transposon integration in human cells. Nucleic Acids Res 2007; 35: e50.

    Article  Google Scholar 

  15. Bowers WJ, Howard DF, Brooks AI, Halterman MW, Federoff HJ . Expression of vhs and VP16 during HSV-1 helper virus-free amplicon packaging enhances titers. Gene Therapy 2001; 8: 111–120.

    Article  CAS  Google Scholar 

  16. Luo G, Ivics Z, Izsvak Z, Bradley A . Chromosomal transposition of a Tc1/mariner-like element in mouse embryonic stem cells. Proc Natl Acad Sci USA 1998; 95: 10769–10773.

    Article  CAS  Google Scholar 

  17. Zayed H, Izsvák Z, Khare D, Heinemann U, Ivics Z . The DNA-bending protein HMGB1 is a cellular cofactor of Sleeping Beauty transposition. Nucleic Acids Res 2003; 31: 2313–2322.

    Article  CAS  Google Scholar 

  18. Yant SR, Kay MA . Nonhomologous-end-joining factors regulate DNA repair fidelity during Sleeping Beauty element transposition in mammalian cells. Mol Cell Biol 2003; 23: 8505–8518.

    Article  CAS  Google Scholar 

  19. Izsvak Z, Stüwe EE, Fiedler D, Katzer A, Jeggo PA, Ivics Z . Healing the wounds inflicted by sleeping beauty transposition by double-strand break repair in mammalian somatic cells. Mol Cell 2004; 13: 279–290.

    Article  CAS  Google Scholar 

  20. Walisko O, Izsvák Z, Szabó K, Kaufman CD, Herold S, Ivics Z . Sleeping Beauty transposase modulates cell-cycle progression through interaction with Miz-1. Proc Natl Acad Sci USA 2006; 103: 4062–4067.

    Article  CAS  Google Scholar 

  21. Yusa K, Takeda J, Horie K . Enhancement of Sleeping Beauty transposition by CpG methylation: possible role of heterochromatin formation. Mol Cell Biol 2004; 24: 4004–4018.

    Article  CAS  Google Scholar 

  22. Mates L, Chuah MK, Belay E, Jerchow B, Manoj N, Acosta-Sanchez A, Grzela DP . Molecular evolution of a novel hyperactive Sleeping Beauty transposase enables robust stable gene transfer in vertebrates. Nat Genet 2009; 41: 753–761.

    Article  CAS  Google Scholar 

  23. Cui Z, Geurts AM, Liu G, Kaufman CD, Hackett PB . Structure-function analysis of the inverted terminal repeats of the sleeping beauty transposon. J Mol Biol 2002; 318: 1221–1235.

    Article  CAS  Google Scholar 

  24. Yant SR, Wu X, Huang Y, Garrison B, Burgess SM, Kay MA et al. High-resolution genome-wide mapping of transposon integration in mammals. Mol Cell Biol 2005; 25: 2085–2094.

    Article  CAS  Google Scholar 

  25. Largaespada DA, Collier LS . Transposon-mediated mutagenesis in somatic cells: identification of transposon-genomic DNA junctions. Methods Mol Biol 2008; 435: 95–108.

    Article  CAS  Google Scholar 

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Acknowledgements

We thank Dr Brendan Lee (Baylor College of Medicine) for providing the DNA spacer elements. NIH U54-NS045309 (HJF/WJB) supported this work.

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Authors and Affiliations

  1. Departments of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA

    S de Silva

  2. Center for Neural Development and Disease, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA

    S de Silva, M A Mastrangelo, L T Lotta Jr, C A Burris & W J Bowers

  3. Departments of Neurology and Neuroscience, Georgetown University Medical Center, Washington, DC, USA

    H J Federoff

  4. Department of Neurology, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA

    W J Bowers

  5. Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA

    W J Bowers

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  1. S de Silva
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  2. M A Mastrangelo
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  3. L T Lotta Jr
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  4. C A Burris
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  5. H J Federoff
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  6. W J Bowers
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Correspondence to W J Bowers.

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de Silva, S., Mastrangelo, M., Lotta, L. et al. Extending the transposable payload limit of Sleeping Beauty (SB) using the Herpes Simplex Virus (HSV)/SB amplicon-vector platform. Gene Ther 17, 424–431 (2010). https://doi.org/10.1038/gt.2009.144

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