December 2002
Volume 43, Issue 13
ARVO Annual Meeting Abstract  |   December 2002
Intercellular spread of HSV-VP22 linked proteins from the Retinal Pigment Epithelium to Photoreceptors
Author Affiliations & Notes
  • R Kumar-Singh
    Ophthalmology and Human Genetics University of Utah Salt Lake City UT
  • S Sadowski
    Salt Lake City UT
  • D Morris
    Salt Lake City UT
  • S Cashman
    Salt Lake City UT
  • Footnotes
    Commercial Relationships   R. Kumar-Singh, None; S. Sadowski, None; D. Morris, None; S. Cashman, None. Grant Identification: Support: Research To Prevent Blindness (RPB), Foundation Fighting Blindness (FFB)
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 2890. doi:
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    • Get Citation

      R Kumar-Singh, S Sadowski, D Morris, S Cashman; Intercellular spread of HSV-VP22 linked proteins from the Retinal Pigment Epithelium to Photoreceptors . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2890.

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

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Abstract: : Purpose: We have determined that Adenovirus 5 (Ad5) mediated gene transfer to photoreceptors is hampered by the lack of Coxsackievirus B-adenovirus Receptors (CAR) on those cells. However, 80% of the cells in the adjacent Retinal Pigment Epithelium (RPE) can be infected by a single injection of Ad5 vectors into the subretinal space. In order to enhance delivery of proteins to photoreceptors by Ad5, we have explored the possibility of using the RPE as a source of therapeutic protein for photoreceptors. Methods: We hypothesized that green fluorescent protein (GFP) fused to the protein transduction domain of Herpes Simplex Virus (HSV) VP22 and delivered by an Ad5 vector to the RPE should allow transfer of the protein from the RPE to the photoreceptors and neurons of the retina. First, we examined intercellular delivery of VP-22 linked GFP in vitro. We transfected ChangC cells with a plasmid co-expressing Vp22-GFP and a Red Fluorescent Protein (RFP). FACS analyses indicated a 32% increase in GFP-only positive cells in comparison to RFP-positive cells. When the same experiment was performed using an Ad5 virus, we determined that there was an increase of 50% GFP-only positive cells compared to the RFP-positive cells. Using Ad5 hexon and DNA Binding Protein antibodies we were able to directly visualize cells that were not infected by virus yet were positive for GFP. We next examined the effects of VP22-GFP in vivo in the context of an Ad5 virus. We injected the sub-retinal space of the right-eye of two month-old C57Bl6 mice with a cmv-Vp22GFPAd5 virus. The left eye was injected with a cmv-GFPAd5 virus. Results: Significant infection of the RPE was observed and a large number of cells in the neural retina including photoreceptors were GFP-positive in Vp22-injected mice, indicating a transfer of GFP from the RPE to those cells. No transfer of GFP from the RPE to neural retina was observed in cmv-GFP injected retina. Conclusions: Therapeutic proteins linked to the protein transduction domain of HSV-VP22 may provide a strategy for efficiently delivering therapeutic proteins to photoreceptors.

Keywords: 419 gene transfer/gene therapy • 568 retinitis • 307 adenovirus 

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