June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Rod outer segments influence the efficiency of AAV-mediated rod transduction
Author Affiliations & Notes
  • Lolita Petit
    Gene Therapy Center / Ophthalmology, University of Massachusetts Medical School, Worcester, Massachusetts, United States
  • Shan Ma
    Gene Therapy Center / Ophthalmology, University of Massachusetts Medical School, Worcester, Massachusetts, United States
  • Shun-Yun Cheng
    Gene Therapy Center / Ophthalmology, University of Massachusetts Medical School, Worcester, Massachusetts, United States
  • Guangping Gao
    Gene Therapy Center / Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, United States
  • Claudio Punzo
    Gene Therapy Center / Ophthalmology / Neurobiology, University of Massachusetts Medical School, Worcester, Massachusetts, United States
  • Footnotes
    Commercial Relationships   Lolita Petit, None; Shan Ma, None; Shun-Yun Cheng, None; Guangping Gao, Voyager Therapeutics (I); Claudio Punzo, None
  • Footnotes
    Support  Information Recherche Retinite Pigmentaire Unrestricted Grant (L.P.), Fulbright/Fondation Monahan Postdoctoral Fellowship (L.P.), Young Researcher in Ophthalmology Postdoctoral Fellowship (L.P.), AFM-Telethon Postdoctoral Fellowship (L.P.), NH Grant EY023570 (C.P.)
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 4094. doi:
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    • Get Citation

      Lolita Petit, Shan Ma, Shun-Yun Cheng, Guangping Gao, Claudio Punzo; Rod outer segments influence the efficiency of AAV-mediated rod transduction. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4094.

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

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Abstract

Purpose : Photoreceptors (PRs) are the primary cells affected in many human retinal diseases. Successfully treating these diseases with gene therapy relies on the efficient transduction of PRs, which remains frequently sub-optimal. PR transduction could be vastly improved by a deeper understanding of the factors that limit PR permissivity to adeno-associated virus (AAV). Interestingly, it has been noted recently that subretinal injection of AAV8 at birth results primarily in cone transduction, whereas the vast majority of transduced cells in adults are rods. Here, we aimed at better understanding this shift in tropism from cones to rods.

Methods : First, CD1 mice were subretinally injected at postnatal (PN) day 1, 5, 10 and 21 with different AAV vectors to identify differences in rod and cone transduction. Experiments used AAV type 5 vectors carrying different transgenes (dGFP, H2bGFP eGFP and Cre) and promoters (CMV, qCMV and mCAR), as well as a panel of 13 AAV serotypes (1, 2, 3, 4, 5, 6, 7, 8, 9, rh8, rh10, rh39 and rh48) all expressing eGFP under the control of the CB6 promoter. Three weeks after delivery, retinal sections and flatmounts were imaged for direct transgene expression using fluorescent microscopy. Immunocytochemistry for cone-specific markers was used to determine the percentage of transduced cones and rods. Second, we evaluated if changes in tropism during PR development correlate with changes in inter-PR matrix binding. This mechanism was tested on PN1 and PN21 retinal explants. Finally, we evaluated if the presence of rod segments influences the tropism using two mouse models of rod-cone dystrophies.

Results : The data revealed that rod transduction temporally and spatially correlates with the development of their segments. All AAVs had an initial preference to transduce cone PRs when rod outer segments are not formed yet (PN1 and PN5), and then displayed a preference for rods as the cells maturate. This shift in tropism did not correlate with changes in AAV binding to the matrix. In contrast, altered development of rod segments was associated with a strong reduction of rod transduction and an increase in the percentage of transduced cones by 2-3.8 fold.

Conclusions : Our results indicate that rod outer segments are necessary for efficient rod transduction and establish access of AAV vectors to rods as one of the major regulatory factors for PR transduction in the adult retina upon subretinal delivery.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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