April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Rpe65-Gene Transfer Using an Integration-Deficient Lentiviral Vector
Author Affiliations & Notes
  • C. Kostic
    Unit of Gene Therapy and Stem Cell Biology, Jules-Gonin Eye Hosp,University Lausanne, Lausanne, Switzerland
  • S. Philippe
    Unit of Gene Therapy and Stem Cell Biology, Jules-Gonin Eye Hosp,University Lausanne, Lausanne, Switzerland
  • S. Crippa
    Unit of Gene Therapy and Stem Cell Biology, Jules-Gonin Eye Hosp,University Lausanne, Lausanne, Switzerland
  • M. Samardzija
    Lab for Retinal Cell Biology, Dept Ophthalmology, University of Zürich, Zürich, Switzerland
  • V. Pignat
    Unit of Gene Therapy and Stem Cell Biology, Jules-Gonin Eye Hosp,University Lausanne, Lausanne, Switzerland
  • D. Wanner
    Unit of Gene Therapy and Stem Cell Biology, Jules-Gonin Eye Hosp,University Lausanne, Lausanne, Switzerland
  • C. Grimm
    Lab for Retinal Cell Biology, Dept Ophthalmology, University of Zürich, Zürich, Switzerland
  • C. Sarkis
    Team of Biotherapy and Biotechnology, CRICM, Paris, France
  • J. Mallet
    Team of Biotherapy and Biotechnology, CRICM, Paris, France
  • Y. Arsenijevic
    Unit of Gene Therapy and Stem Cell Biology, Jules-Gonin Eye Hosp,University Lausanne, Lausanne, Switzerland
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 4498. doi:
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      C. Kostic, S. Philippe, S. Crippa, M. Samardzija, V. Pignat, D. Wanner, C. Grimm, C. Sarkis, J. Mallet, Y. Arsenijevic; Rpe65-Gene Transfer Using an Integration-Deficient Lentiviral Vector. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4498.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : We previously demonstrated efficient retinal rescue of RPE65 mouse models (Rpe65-/- (Bemelmans et al, 2006) and Rpe65R91W/R91W mice) using a HIV1-derived lentiviral vector encoding for the mouse RPE65 cDNA. In order to optimize a lentiviral vector as an alternative tool for RPE65-derived Leber Congenital Amaurosis clinical trials, we evaluated the efficiency of an integration-deficient lentiviral vector (IDLV) encoding the human RPE65 cDNA to restore retinal function in the Rpe65R91W/R91W mice.

Methods: : An HIV-1-derived lentiviral vector expressing either the hrGFPII or the human Rpe65 cDNA under the control of a 0.8 kb fragment of the human Rpe65 promoter (R0.8) was produced by transient transfection of 293T cells. A LQ-integrase mutant was used to generate the IDLV vectors. IDLV-R0.8-hRPE65 or hrGFPII were injected subretinally into 1 month-old Rpe65R91W/R91W mice. Functional rescue was assessed by ERG (1 and 3 months post-injection) and cone survival by immunohistology.

Results: : An increased light sensitivity was detected by scotopic ERG in animals injected with IDLV-R0.8-hRPE65 compared to hrGFPII-treated animals or untreated mice. However the improvement was delayed compared to integration-proficient LV and observed at 3 months but not 1 month post-injection. Immunolabelling of cone markers showed an increased number of cones in the transduced area compared to control groups.

Conclusions: : The IDLV-R0.8-hRPE65 vectors allow retinal improvement in the Rpe65R91W/R91W mice. Both rod function and cone survival were demonstrated even if there is a delay in the rescue as assessed by scotopic ERG. Integration-deficient vectors minimize insertional mutagenesis and thus are safer candidates for human application. Further experiments using large animals are now needed to validate correct gene transfer and expression of the RPE65 gene as well as tolerance of the vector after subretinal injection before envisaging a clinical trial application.

Keywords: gene transfer/gene therapy • retinal degenerations: hereditary • retinal pigment epithelium 
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