March 2012
Volume 53, Issue 14
Free
ARVO Annual Meeting Abstract  |   March 2012
Evaluation Of AAV-DJ Vector As A Therapeutic Delivery System For Ocular Cells And Tissue
Author Affiliations & Notes
  • Matthew Hartzell
    Opthalmology, Oregon Health and Science University, Portland, Oregon
  • Maria Parker
    Opthalmology, Oregon Health and Science University, Portland, Oregon
  • Andrew Stempel
    Opthalmology, Oregon Health and Science University, Portland, Oregon
  • Trevor McFarland
    Opthalmology, Oregon Health and Science University, Portland, Oregon
  • Binoy Appukuttan
    Opthalmology, Oregon Health and Science University, Portland, Oregon
  • John T. Stout
    Opthalmology, Oregon Health and Science University, Portland, Oregon
  • Footnotes
    Commercial Relationships  Matthew Hartzell, None; Maria Parker, None; Andrew Stempel, None; Trevor McFarland, None; Binoy Appukuttan, None; John T. Stout, None
  • Footnotes
    Support  NIH;NEI;RO1 EY019042
Investigative Ophthalmology & Visual Science March 2012, Vol.53, 1895. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Matthew Hartzell, Maria Parker, Andrew Stempel, Trevor McFarland, Binoy Appukuttan, John T. Stout; Evaluation Of AAV-DJ Vector As A Therapeutic Delivery System For Ocular Cells And Tissue. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1895.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose: : Gene therapy for the treatment of Leber Congenital Amerosis (LCA) using an AAV vector is currently under a Phase I trial. The development of adeno-associated viral (AAV) vectors that can deliver to multiple ocular cell types with high efficiency via subretinal or intravitreal delivery methods is essential for effective therapy. The AAV-DJ (type2/type8/type9 chimera) was engineered from shuffling 8 different wild type native viruses (Kay et al 2008). The goal of this study is to investigate whether the chimeric AAV-DJ-GFP vector is capable of transducing a variety of ocular cell types in vitro and in vivo.

Methods: : The chimeric AAV- DJ vector serotype (AAV-DJ) encoding enhanced green fluorescent protein (GFP) was produced (Vollum Viral Core). One microliter of virus with a titer of 4x10e12 GC/ml was added to the following cell lines: HEK 293T, mouse Muller glia (C57MIO), human Muller glia (MIO-M7), primary human and primary pig trabecular meshwork, human retinal pigment epithelial (ARPE 19), rhesus retinal/choroidal endothelial (RF/6A), human primary iris endothelial /fibroblast, retinoblastoma cells (Y79), human choroidal endothelial and ocular melanoma (Mel 202). Cells were cultured and GFP expression was measured by fluorescent microscopy at 18 and 42 hours.1.5ul of AAV-DJ was injected intravitreally in each eye of a C57BL/6 mouse. Mice were euthanized on post-injection day 5. Retinal flat mounts were prepared. GFP -positive cells were viewed using fluorescent microscopy.

Results: : With the exception of Y79 retinoblastoma cells, all cell types cultured were transduced with high efficiency and GFP expression was observed at all time points. Multiple GFP positive cells were observed within the neural retina after intravitreal injection.

Conclusions: : The 2-8-9 chimera AAV-DJ vector can transduce multiple ocular derived cells in culture. The AAV-DJ vector has the potential to be useful for ocular cell studies as well as gene therapy experiments.

Keywords: gene transfer/gene therapy 
×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×