Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Recombinant adeno-associated virus for muscle directed gene therapy

Nature Medicine volume 3, pages 306–312 (1997)Cite this article

Abstract

Although gene transfer with adeno-associated virus (AAV) vectors has typically been low, transduction can be enhanced in the presence of adenovirus gene products through the formation of double stranded, non-integrated AAV genomes. We describe the unexpected finding of high level and stable transgene expression in mice following intramuscular injection of purified recombinant AAV (rAAV). The rAAV genome is efficiently incorporated into nuclei of differentiated muscle fibers where it persists as head-to-tail concatamers. Fluorescent in situ hybridization of muscle tissue suggests single integration sites. Neutralizing antibody against AAV capsid proteins does not prevent readministration of vector. Remarkably, no humoral or cellular immune responses are elicited to the neoantigenic transgene product E. coli β-galactosidase. The favorable biology of rAAV in muscle-directed gene therapy described in this study expands the potential of this vector for the treatment of inherited and acquired diseases.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Kotin, R.M. Prospects for the use of adeno-associated virus as a vector for human gene therapy. Hum. Gene Ther. 5, 793–801 (1994).

    Article  CAS  PubMed  Google Scholar 

  2. Kotin, R. et al. Site-specific integration by adeno-associated virus. Proc. Natl. Acad. Sci. USA 87, 2211–2215 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Samulski, R. et al. Targeted integration of adeno-associated virus (AAV) into human chromosome 19. EMBO J. 10, 3941–3950 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Berns, K.I. Parvovirus replication. Microbiol. Rev. 54, 316–329 (1990).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Samulski, R., Chang, L. & Shenk, T. Helper-free stocks of recombinant adeno-associated viruses: Normal integration does not require viral gene expression. J. Virol. 63, 3822–3828 (1989).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Russell, D., Miller, A. & Alexander, I. Adeno-associated virus vectors preferentially transduce cells in S phase. Proc. Natl. Acad. Sci. USA 91, 8915–8919 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Alexander, I.E., Russell, D.W. & Miller, A.D. DNA-damaging agents greatly increase the transduction of nondividing cells by adeno-associated virus vectors. J. Virol. 68, 8282–8287 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Russell, D.W., Alexander, I.E. & Miller, A.D. DNA synthesis and topoisomerase inhibitors increase transduction by adeno-associated virus vectors. Proc. Natl. Acad. Sci. USA 92, 5719–5723 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Fisher, K.J. et al. Transduction with recombinant adeno-associated virus for gene therapy is limited by leading-strand synthesis. J. Virol. 70, 520–532 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Ferrari, F.K., Samulski, T., Shenk, T. & Samulski, R.J. Second-strand synthesis is a rate-limiting step for efficient transduction by recombinant adeno-associated virus vectors. J. Virol. 70, 3227–3234 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Kumar, S. & Leffak, M. Conserved chromatin structure in c-myc 5′-flanking DNA after viral transduction. J. Mol. Biol. 222, 45–57 (1991).

    Article  CAS  PubMed  Google Scholar 

  12. Kaplitt, M.G. et al. Long-term gene expression and phenotypic correction using adenoassociated virus vectors in the mammalian brain. Nature Genet. 8, 148–154 (1994).

    Article  CAS  PubMed  Google Scholar 

  13. Miller, J.L. et al. Recombinant adeno-associated virus (rAAV)-mediated expression of a human g-globin gene in human progenitor-derived erythroid cells. Proc. Natl. Acad. Sci. USA 91, 10183–10187 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Podsakoff G., Wong K.K., Jr. & Chatterjee, S. Efficient gene transfer into nondividing cells by adeno-associated virus-based vectors. J. Virol. 68, 5656–5666 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Walsh, C.E. et al. Phenotypic correction of Fanconi anemia in human hematopoietic cells with a recombinant adeno-associated virus vector. J. Clin. Invest. 94, 1440–1448 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Flotte, T.R. et al. Stable in vivo expression of the cystic fibrosis transmembrane conductance regulator with an adeno-associated virus vector. Proc. Natl. Acad. Sci. USA 90, 10613–10617 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Tratschin, J.-D., Miller, I.L., Smith, M.G. & Carter, B.J. Adeno-associated virus vector for high-frequency integration, expression, and rescue of genes in mammalian cells. Mol. Cell. Biol. 5, 3251–3260 (1985).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Muzcyzka, N. Use of adeno-associated virus as a general transduction vector in mammalian cells. Curr. TopicsMicrobiol. Immunol. 158, 97–129 (1992).

    Google Scholar 

  19. McLaughlin, S.K., Collis, P., Hermonat, P.L. & Muzyczka, N. Adeno-associated virus general transduction vectors: Analysis of proviral structures. J. Virol. 62, 1963–1973 (1988).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Yang, Y., Haecker, S.E., Su, Q. & Wilson, J.M. Immunology of gene therapy with adenoviral vectors in the mouse skeletal muscle. Hum. Molec. Genet. 5, 1703–1712 (1996).

    Article  CAS  PubMed  Google Scholar 

  21. Trask, B.J. Fluorescence in situ hybridization applications in cytogenesis and genemapping. Trends Genet. 7, 149–154 (1991).

    Article  CAS  PubMed  Google Scholar 

  22. Blau, H.M., Pavlath, G.K., Rich, K. & Webster, S.G. Localization of muscle gene products in nuclear domains: Does this constitute a problem for myoblast therapy? Adv. Exp. Med. Biol. 280, 167–172 (1990).

    Article  CAS  PubMed  Google Scholar 

  23. Howe, J.R., Skryabin, B.V., Belcher, S.M., Zerillo, C.A. & Schmauss, C. The responsiveness of a tetracycline-sensitive expression system differs in different cell lines. J. Biol. Chem. 270, 14168–14174 (1995).

    Article  CAS  PubMed  Google Scholar 

  24. Rivera, V.M. et al. A humanized system for pharmacologic control of gene expression. Nature Med. 2, 1028–1032 (1996).

    Article  CAS  PubMed  Google Scholar 

  25. Wolff, J.A. et al. Direct gene transfer into mouse muscle in vivo. Science 247, 1465–1468 (1990).

    Article  CAS  PubMed  Google Scholar 

  26. Tripathy, S.K., Black, H.B., Goldwasser, E. & Leiden, J.M. Immune responses to transgeneencoded proteins limit the stability of gene expression after injection of replication-defective adenovirus vectors. Nature Med. 2, 545–550 (1996).

    Article  CAS  PubMed  Google Scholar 

  27. Dhawan, J. et al. Systemic delivery of human growth hormone by injection of genetically engineered myoblasts. Science 254, 1509–1512 (1991).

    Article  CAS  PubMed  Google Scholar 

  28. Barr, E. & Leiden, J.M. Systemic delivery of recombinant proteins by genetically modified myoblasts. Science 254, 1507–1509 (1991).

    Article  CAS  PubMed  Google Scholar 

  29. Yao, S.-N., Smith, K.J. & Kurachi, K. Primary myoblast-mediated gene transfer: Persisitent expression of human factor IX in mice. Gene Ther. 1, 99–107 (1994).

    CAS  PubMed  Google Scholar 

  30. Gussoni, E. et al. A method to co-detect introduced genes and their products in gene therapy protocols. Nature Biotech. 14, 1012–1016 (1996).

    Article  CAS  Google Scholar 

  31. Bradford, M.M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254 (1976).

    Article  CAS  PubMed  Google Scholar 

  32. Rice, S.A. & Klessig, D.F. Isolation and analysis of adenovirus type 5 mutants containing deletions in the gene encoding the DNA binding protein. J. Virol. 56, 767–778 (1985).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Wolfe, J.H. et al. Restoration of normal lysosomal function in mucopolysaccharidosis type VII cells by retroviral vector-mediated gene transfer. Proc. Natl. Acad. Sci. USA 87, 2877–81 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Institute for Human Gene Therapy and Departments of Medicine and Molecular and Cellular Engineering,

    Krishna J. Fisher, Krishna J. Fisher, Karin Jooss, Karin Jooss, James Alston, James Alston, Yiping Yang, Yiping Yang, Sarah Ehlen Haecker, Sarah Ehlen Haecker, Katherine High, Ravindra Pathak, Ravindra Pathak, Steven E. Raper, James M. Wilson & James M. Wilson

  2. Surgery University of Pennsylvania Health System, Philadelphia, Pennsylvania, 19104, USA

    Steven E. Raper

  3. Pediatrics University of Pennsylvania Health System, Philadelphia, Pennsylvania, 19104, USA

    Katherine High

  4. The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, 19104, USA

    Katherine High

  5. The Wistar Institute, 3601 Spruce Street, Philadelphia, Pennsylvania, 19104, USA

    James M. Wilson

Authors
  1. Krishna J. Fisher
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Karin Jooss
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. James Alston
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yiping Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sarah Ehlen Haecker
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Katherine High
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ravindra Pathak
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Steven E. Raper
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. James M. Wilson
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Fisher, K., Jooss, K., Alston, J. et al. Recombinant adeno-associated virus for muscle directed gene therapy. Nat Med 3, 306–312 (1997). https://doi.org/10.1038/nm0397-306

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm0397-306

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing