June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Optimization of Cone-Directed AAV-Mediated Gene Augmentation Therapy for CNGB3-Achromatopsia by Use of the IRBP/GNAT2-Promoter and Intravitreal CNTF Administration
Author Affiliations & Notes
  • Connie Yeh
    School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA
    College of Veterinary Medicine, Michigan State University, East Lansing, MI
  • Simone Iwabe
    School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA
  • Sanford Boye
    College of Medicine, University of Florida, Gainesville, FL
  • Kendra McDaid
    School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA
  • Christine Harman
    College of Veterinary Medicine, Michigan State University, East Lansing, MI
  • Rong Wen
    Bascom Palmer Eye Institute, University of Miami, Miami, FL
  • William Hauswirth
    College of Medicine, University of Florida, Gainesville, FL
  • Andras Komaromy
    School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA
    College of Veterinary Medicine, Michigan State University, East Lansing, MI
  • Gustavo Aguirre
    School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA
  • Footnotes
    Commercial Relationships Connie Yeh, None; Simone Iwabe, None; Sanford Boye, PCT/US2012/062478 (P); Kendra McDaid, None; Christine Harman, None; Rong Wen, Neurotech USA (C); William Hauswirth, AGTC (I), Bionic Sight (I), AGTC (C), Syncona (C), RetroSense (C); Andras Komaromy, None; Gustavo Aguirre, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 1937. doi:
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      Connie Yeh, Simone Iwabe, Sanford Boye, Kendra McDaid, Christine Harman, Rong Wen, William Hauswirth, Andras Komaromy, Gustavo Aguirre; Optimization of Cone-Directed AAV-Mediated Gene Augmentation Therapy for CNGB3-Achromatopsia by Use of the IRBP/GNAT2-Promoter and Intravitreal CNTF Administration. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1937.

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

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Abstract

Purpose: AAV5-mediated cone-directed gene therapy results in rescue of cone function and day vision in canine models of CNGB3-achromatopsia. However, the treatment success rate is <80%, and the human red-cone opsin promoter does not target S-cones. The objective of this study was to optimize therapeutic outcome (1) by use of an enhanced cone transducin promoter and (2) by intravitreal CNTF administration in non-responders.

Methods: The IRBP/GNAT2 promoter consists of the 277-bp 5'-flanking sequence of the human GNAT2 promoter coupled with the 214-bp IRBP enhancer (Ying et al. 1998). The ability of this promoter to target cone subclasses was evaluated by subretinal injection of AAV5-IRBP/GNAT2-GFP in normal dog eyes. GFP reporter gene expression was evaluated by observation of in vivo fluorescence and by immunohistochemistry. Subsequently, 14 young CNGB3-mutant dogs between 9-11 weeks of age were treated by bilateral subretinal injection of either AAV5-IRBP/GNAT2-hCNGB3 or AAV8-IRBP/GNAT2-hCNGB3. The animals were followed for 5-15 months post injection by standard full-field electroretinography and visual behavior testing in an obstacle avoidance course. Six months post therapy, 3 “non-responder” dogs were intravitreally injected with 12 μg of CNTF (n = 3 eyes) or PBS (n = 3 eyes) and followed for 2 months.

Results: The IRBP/GNAT2 promoter allowed robust and specific targeting of GFP-reporter gene expression in both cone subclasses. Rescue of cone function and day vision was achieved with both AAV serotypes; best results were obtained by use of ~1012 vector genomes/mL (2/6 and 4/6 injected eyes responding with AAV5 and AAV8 respectively). A therapeutic effect was not obtained in all AAV-treated eyes. However, all 3 eyes treated with intravitreal CNTF 6 months following unsuccessful gene augmentation therapy showed robust rescue of cone function; this was not the case with intravitreal PBS injection.

Conclusions: Gene expression can be targeted to both canine cone subclasses and their function restored in CNGB3-achromatopsia with the IRBP/GNAT2 promoter in either AAV5 or AAV8. It remains unclear why some animals do not respond to AAV-mediated gene replacement therapy alone; however, treatment outcome can be enhanced by combination with intravitreal CNTF injection in eyes that do not respond.

Keywords: 648 photoreceptors • 490 cytokines/chemokines • 503 drug toxicity/drug effects  
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