May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
A Novel Prostanoid Pathway-Based Lentiviral Vector Gene Therapy System to Reduce Intraocular Pressure
Author Affiliations & Notes
  • R. A. Barraza
    Molecular Medicine Program, Mayo Clinic College of Medicine, Rochester, Minnesota
  • E. M. Poeschla
    Molecular Medicine Program, Mayo Clinic College of Medicine, Rochester, Minnesota
  • Footnotes
    Commercial Relationships  R.A. Barraza, None; E.M. Poeschla, None.
  • Footnotes
    Support  NIH Grant EY01141
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 2620. doi:https://doi.org/
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      R. A. Barraza, E. M. Poeschla; A Novel Prostanoid Pathway-Based Lentiviral Vector Gene Therapy System to Reduce Intraocular Pressure. Invest. Ophthalmol. Vis. Sci. 2008;49(13):2620. doi: https://doi.org/.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose: : We previously introduced a novel cyclooxygenase-2 (COX-2) and prostaglandin (PG)-based gene therapy system that significantly reduces intraocular pressure (IOP) in a sustained manner. A single concomitant delivery of COX-2 and FPR (PG F receptor) is sufficient for sustained IOP reduction. In this system, codon optimization (co) of COX-2 cDNA was required for robust PG synthesis. We further developed new vectors for this therapeutic system, including a single bicistronic vector expressing all required transgenes.

Methods: : RT Q-PCR determined RNA decay rates, and RNA in situ hybridization was performed in coCOX-2- and wtCOX-2-transduced cells. We engineered and validated new COX2iPGFS (PGF synthase) and COX2iFPR vectors by Western blot, PGF EIA, and a Nur77 promoter-dependent luciferase reporter assay. We transcorneally injected 107 TU FPRiG, COX2iG with FPRiG, COX2iPGFS with FPRiG, or COX2iFPR in right eyes. ACs of 19 cats (38 eyes) were injected, with L eyes receiving 107 TU control vector (intra-animal control design). IOP was monitored serially.

Results: : In situ hybridization revealed striking nuclear sequestration of wtCOX-2 mRNA compared to cytoplasmic localization of coCOX-2 mRNA. Increased message stability was observed for coCOX-2 compared to wtCOX-2. We confirmed FPR activity with a PGF dose-dependent increase in reporter activity. EIA confirmed COX-2 and PGFS activity. Concomitant COX2iPGFS and FPRiG administration resulted in sustained IOP reduction.

Conclusions: : Codon-optimization of COX-2 confers mRNA stability and nuclear export, leading to robust expression. Single lentivectors expressing both functional COX-2 and FPR, or COX-2 and PGFS function well. Animal studies of COX2iFPR and the effects of pharmacologic inhibitors and agonists on FPR-expressing cat TMs are currently underway.

Keywords: gene transfer/gene therapy • intraocular pressure • anterior chamber 
×
×

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.

×