Advertisement

Ribozyme-Mediated Gene Therapy for Autosomal Dominant Retinal Degeneration

  • John G. Flannery
  • Alfred S. Lewin
  • Kimberly A. Drenser
  • Shimpei Nishikawa
  • Douglas Yasumura
  • Matthew M. LaVail
  • William W. Hauswirth

Abstract

Ribozymes targeted to a point mutation (histidine for proline substitution at position 23) in rhodopsin were evaluated as therapeutic agents for photoreceptor degeneration in an animal model of autosomal dominant retinitis pigmentosa (ADRP). Recombinant adeno-associated virus (AAV) vectors incorporating a proximal bovine rod opsin promoter were used to transfer either hairpin or hammerhead ribozyme genes to photoreceptors. AAV was administered by subretinal injection at postnatal day (P) 15 to rats expressing a rod opsin transgene with a proline-to-histidine substitution in exon 1 at codon 23 of the rhodopsin gene (P23H). In vivo levels of normal and mutant RNAs were measured by allele-specific RT-PCR at P60. Eyes were examined by histopathology and morphometric analysis, as well as by ERG at approximately P60, P75 or P90. Expression of either a hammerhead or hairpin ribozyme markedly slowed the rate of photoreceptor degeneration for at least 3 months as determined by outer nuclear layer thickness as well as by inner and outer segment length. Catalytically inactive control ribozymes had a significantly smaller effect on the retinal degeneration. Minimal rescue effects were seen in retinas injected with either AAV containing a gene for green fluorescent protein rather than the ribozyme gene or buffer only. The level of mutant opsin RNA relative to wild-type RNA was also reproducibly lower in ribozyme-treated retina. Finally, ERG comparisons of ribozyme-treated versus control eyes demonstrated functional rescue. We conclude that ribozyme-directed cleavage of mutant mRNAs appears to be a potentially effective therapy for autosomal dominant retinal degeneration.

Keywords

Retinal Degeneration Outer Nuclear Layer Hammerhead Ribozyme Photoreceptor Degeneration Autosomal Dominant Retinitis Pigmentosa 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    P. Steinecke, T. Herget, and P.H. Schreier, 1992, Expression of a chimeric ribozyme gene results in endonucleolytic cleavage of target mRNA and a concomitant reduction of gene expression in vivo., EMBO. J. 11:1525–1530.PubMedGoogle Scholar
  2. 2.
    K.R. Birikh, P.A. Heaton, and F. Eckstein, 1997, The structure, function, and application of the hammerhead ribozyme., Eur. J. Biochem. 245:1–16.PubMedCrossRefGoogle Scholar
  3. 3.
    J.J. Rossi, 1997, Therapeutic applications of catalytic antisense RNAs (ribozymes)., Ciba. Found. Symp. 209. 195–204.PubMedGoogle Scholar
  4. 4.
    ML Cotten and M.L. Birnstiel, 1989, Ribozyme mediated destruction of RNA in vivo., EMBO. J. 8:3861–3866.PubMedGoogle Scholar
  5. 5.
    A. Lieber, and M.A. Kay, 1996, Adenovirus-mediated expression of ribozymes in mice., J. Virol. 70:3153–3158.PubMedGoogle Scholar
  6. 6.
    P. L’Huillier, S. Soulier, and M.G. Stinnakre, et al., 1996, Efficient and specific ribozyme-mediated reduction of bovine alpha-lactalbumin expression in double transgenic mice., Proc. Natl. Acad. Sci. 93:6698–6703.PubMedCrossRefGoogle Scholar
  7. 7.
    X. Xiao, J. Li, and R.J. Samulski, 1996, Efficient long-term transfer into muscle tissue of immuno-competent mice by adeno-associated virus vector., J. Virol. 70:8098–8108.PubMedGoogle Scholar
  8. 8.
    K.J. Fisher, K. Jooss, and J. Alston, et al., 1997, Recombinant adeno-associated virus for muscle directed gene therapy., Nat. Medicine. 3:306–312.CrossRefGoogle Scholar
  9. 9.
    J.G. Flannery, S. Zolotukhin, M.I. Vaquero, M.M. LaVail, N. Muzyczka, and W.W. Hauswirth, 1997, Efficient photoreceptor-targeted gene expression in vivo by recombinant adeno-associated virus. Proceedings of the National Academy of Sciences of the United States of America. 94(13):6916–6921.PubMedCrossRefGoogle Scholar
  10. 10.
    S. Zolotukhin, M. Potter, W.W. Hauswirth, J. Guy, and N. Muzyczka, 1996, A “humanized” green fluorescent protein cDNA adapted for high-level expression in mammalian cells., J. Virol. 70:4646–4654.PubMedGoogle Scholar
  11. 11.
    L.E. DesJardin and W.W. Hauswirth, 1996, Developmentally important DNA elements within the bovine opsin upstream region, Investigative Ophthalmology and Visual Science. 37(1):154–165.PubMedGoogle Scholar
  12. 12.
    K.A. Drenser, A.M. Timmers, W.W. Hauswirth, and A.S. Lewin, 1998, Ribozyme-targeted destruction of RNA associated with autosomal-dominant retinitis pigmentosa, Investigative Ophthalmology and Visual Science. 39(5):681–689.PubMedGoogle Scholar
  13. 13.
    M. Altschuler, R. Tritz, and A. Hampel, 1992, A method for generating transcripts with defined 5’ and 3’ termini by autolytic processing., Gene. 122:85–90 (1992).PubMedCrossRefGoogle Scholar
  14. 14.
    E. Bertrand, D. Castanotto, and C. Zhou, et al., 1997, The expression cassette determines the functional activity of ribozymes in mammalian cells by controlling their intracellular localization., RNA. 3:75–88.PubMedGoogle Scholar
  15. 15.
    T.R. Flotte and B. Carter, 1995, Adeno-associated virus vectors for gene therapy., Gene Therapy. 2:357–362.PubMedGoogle Scholar
  16. 16.
    R.H. Steinberg, J.G. Flannery, M.I. Naash, J. Chen, M. Matthes, D. Yasumura, and M.M. LaVail, 1996, Transgenic rat models of inherited retinal degeneration caused by mutant opsin genes., Inv. Ophth. Vis. Sci. 37. S698.Google Scholar
  17. 17.
    M.M. LaVail and B.A. Battelle, 1975, Influence of eye pigmentation and light deprivation on inherited retinal dystrophy in the rat., Exp. Eye Res. 21:167–192.PubMedCrossRefGoogle Scholar
  18. 18.
    E.G. Faktorovich, R.H. Steinberg, D. Yasumura, M.T. Matthes, and M.M. LaVail. 1990, Photoreceptor degeneration in inherited dystrophy delayed by the basic fibroblast growth factor., Nature. 347:83–86.PubMedCrossRefGoogle Scholar
  19. 19.
    A.M. Timmers, B.R. Newton, and W.W. Hauswirth, 1993, Synthesis and stability of retinal photorecptor mRNAs are coordinately regulated during bovine fetal development., Exp. Eye Res. 56:251–265.CrossRefGoogle Scholar
  20. 20.
    J.J. Michon, Z.L. Li, S.N. Zhioura, R.J. Anderson, and M.O. Tso, 1991, A comparative study of methods of photoreceptor morphometry., Invest. Ophthalmol. Vis. Sci. 32:280–284.PubMedGoogle Scholar
  21. 21.
    R. Wen, Y. Song, and T. Cheng, et al., 1995, Injury-induced upregulation of bFGF and CNTF mRNAS in the rat retina., J. Neumsci. 15:7377–7385.Google Scholar
  22. 22.
    R. Hormes, M. Homann, and I. Oelze, et al., 1997, The subcellular localization and length of hammerhead ribozymes determine efficacy in human cells., Nucleic. Acids. Res. 25:769–775.PubMedCrossRefGoogle Scholar
  23. 23.
    A.S. Lewin, K.A. Drenser, W.W. Hauswirth, S. Nishikawa, D. Yasumura, J.G. Flannery, and M.M. LaVail, 1998, Ribozyme rescue of photoreceptor cells in a transgenic rat model of autosomal dominant retinitis pigmentosa, Nature Medicine. 4(8):967–71.PubMedCrossRefGoogle Scholar
  24. 24.
    N. Mittereder, K.L. March, and B.C. Trapnell, 1996, Evaluation of the concentration and bioactivity of adenovirus vectors for gene therapy., J. Virol. 70:7498–7509.PubMedGoogle Scholar
  25. 25.
    C. Nyberg-Hoffman, P. Shabram, W. Li, D. Giroux, and E. Aguilar-Cordova, 1997, Sensitivity and reproducibility in adenoviral infectious titer determination., Nature Medicine. 3:808–811.PubMedCrossRefGoogle Scholar
  26. 26.
    H. Miyoshi, M. Takahashi, F. Gage, and I. Verma, 1997, Stable and efficient gene transfer into the retina using an HIV-based lentiviral vector, PNAS. 94:10319–10323.PubMedCrossRefGoogle Scholar
  27. 27.
    S. Mohand-Said, D. Hicks, and M. Simonutti, et al., 1997, Photoreceptor transplants increase host cone survival in the retinal degeneration (rd) mouse., Ophthalmic Res. 29:290–297.PubMedCrossRefGoogle Scholar
  28. 28.
    P.C. Huang, A.E. Gaitan, Y. Hao, R.M. Petters, and F. Wong, 1993, Cellular interactions implicated in the mechanism of photoreceptor degeneration in transgenic mice expressing a mutant rhodopsin gene, Proceedings of the National Academy of Sciences of the United States of America. 90(18):8484–8488.PubMedCrossRefGoogle Scholar

Copyright information

© Kluwer Academic / Plenum Publishers 1999

Authors and Affiliations

  • John G. Flannery
    • 1
  • Alfred S. Lewin
    • 2
    • 4
  • Kimberly A. Drenser
    • 2
  • Shimpei Nishikawa
    • 5
  • Douglas Yasumura
    • 5
  • Matthew M. LaVail
    • 5
  • William W. Hauswirth
    • 2
    • 3
    • 4
  1. 1.Departments of Vision Science, Neuroscience, and BiophysicsUniversity of CaliforniaBerkeley
  2. 2.Department of Molecular Genetics and MicrobiologyUniversity of Florida College of MedicineGainesville
  3. 3.Department of OphthalmologyUniversity of Florida College of MedicineGainesville
  4. 4.Gene Therapy CenterUniversity of Florida College of MedicineGainesville
  5. 5.Departments of Anatomy and Ophthalmology Beckman Vision CenterUniversity of California School of MedicineSan Francisco

Personalised recommendations