April 2009
Volume 50, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2009
Treatment of Corneal Neovascularization Through Transposase-Mediated Somatic Insertional Mutagenesis for Long-Term Expression of Flt2-3k
Author Affiliations & Notes
  • M. S. Hansen
    Department of Ophthalmology, University of Utah, Salt Lake City, Utah
  • N. Singh
    Department of Ophthalmology, University of Utah, Salt Lake City, Utah
  • V. Jessop
    Department of Ophthalmology, University of Utah, Salt Lake City, Utah
    Department of Anatomy, Biochemistry, and Physiology, University of Hawaii, Honolulu, Hawaii
  • S. Moisyadi
    Department of Anatomy, Biochemistry, and Physiology, University of Hawaii, Honolulu, Hawaii
  • B. Ambati
    Department of Ophthalmology, University of Utah, Salt Lake City, Utah
  • Footnotes
    Commercial Relationships  M.S. Hansen, None; N. Singh, None; V. Jessop, None; S. Moisyadi, None; B. Ambati, None.
  • Footnotes
    Support  NEI R01 EY017182
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 4958. doi:
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    • Get Citation

      M. S. Hansen, N. Singh, V. Jessop, S. Moisyadi, B. Ambati; Treatment of Corneal Neovascularization Through Transposase-Mediated Somatic Insertional Mutagenesis for Long-Term Expression of Flt2-3k. Invest. Ophthalmol. Vis. Sci. 2009;50(13):4958.

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

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Abstract

Purpose: : To determine whether, FLT2-3K intraceptor, a recombinant construct of domains 2 to 3 of VEGFR-1 (Flt) coupled with an endoplasmic reticulum retention signal (KDEL), can be expressed for long periods using somatic integration.

Methods: : In order to attain long-term expression of FLT2-3K intraceptor, that sequesters VEGF, we have utilized the insertional property of the piggybac transposase. Transposases are enzymes with the ability of mobilizing DNA via a cut-and-paste mechanism. Transposon-mediated mutagenesis has largely been reserved for genetic manipulating of lower organisms because of low integration rates and nonspecific integration, which can lead to undesirable consequences. Piggybac has been shown to have the highest integration efficiencies compared to other transposases. We generated a fusion protein using the piggybac vector that will express FLT2-3K intraceptor for long periods using non-viral somatic genomic integration.

Results: : Preliminary results in vitro showed that the piggybac mediated expression protein proved to be a successful system for specific insertional mutagenesis. Integration of desired DNA was determined by visualization of GFP within the cells.

Keywords: cornea: basic science • gene transfer/gene therapy 
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