June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
Transduction characteristics of single-stranded and self-complementary AAV2.8 in the porcine retina
Author Affiliations & Notes
  • Silja Hansen
    Biomedicine, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Anna Bøgh Lindholm
    Biomedicine, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Thomas Stax Jakobsen
    Biomedicine, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Lars Aagaard
    Biomedicine, Aarhus Universitet, Aarhus, Midtjylland, Denmark
  • Toke Bek
    Department of Ophthalmology, Aarhus Universitetshospital, Aarhus, Denmark
  • Anne Louise Askou
    Biomedicine, Aarhus Universitet, Aarhus, Midtjylland, Denmark
    Department of Ophthalmology, Aarhus Universitetshospital, Aarhus, Denmark
  • Thomas Corydon
    Biomedicine, Aarhus Universitet, Aarhus, Midtjylland, Denmark
    Department of Ophthalmology, Aarhus Universitetshospital, Aarhus, Denmark
  • Footnotes
    Commercial Relationships   Silja Hansen None; Anna Lindholm None; Thomas Jakobsen None; Lars Aagaard None; Toke Bek None; Anne Louise Askou None; Thomas Corydon None
  • Footnotes
    Support  Aarhus University, Fight for Sight Denmark, Synoptik Fonden, Øjenfonden
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 780. doi:
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      Silja Hansen, Anna Bøgh Lindholm, Thomas Stax Jakobsen, Lars Aagaard, Toke Bek, Anne Louise Askou, Thomas Corydon; Transduction characteristics of single-stranded and self-complementary AAV2.8 in the porcine retina. Invest. Ophthalmol. Vis. Sci. 2023;64(8):780.

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

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Abstract

Purpose : The transduction efficacy and retinal safety of self-complementary (sc) AAV2.8 and single-stranded (ss) AAV2.8 remains to be tested in larger preclinical models. Here, we evaluated key parameters of AAV-transduction following subretinal injection in the porcine retina.

Methods : 14 eyes of 7 domestic pigs received a subretinal injection of GFP encoding scAAV2.8 (scAAV2.8-GFP) or ssAAV2.8-GFP at a dose of 1 x 10^10 vg/eye (50 uL). The transduction efficacy was evaluated in vivo using fundoscopy at day 14, 28, and 42 post injection (PI). Retinal integrity was assessed by optical coherence tomography (OCT). For each timepoint, in two eyes per group (scAAV8-GFP and ssAAV8-GFP) GFP expression was evaluated on RPE/choroidal flatmounts. The transduction profile through the retinal layers was assessed on retinal cross-sections. Slit lamp examination and fundoscopy were used to monitor for intraocular inflammation.

Results : A total of 15/16 eyes were included for data analysis. One eye was excluded. For eyes injected with scAAV2.8-GFP in vivo analysis of GFP expression confirmed retinal transduction in 7/7, 5/5, and 3/3 eyes 14, 28, and 42 days PI, respectively. The corresponding numbers for ssAAV2.8-GFP were 3/6, 3/4, and 3/3. Eyes injected with scAAV2.8 initiated expression earlier, which increased at a faster rate, and ultimately leading to higher levels. Preliminary results of fundoscopy in vivo, suggest a peak of GFP expression at day 28 for scAAV2.8 and at day 42 for ssAAV2.8 Ex vivo analysis showed GFP expression for both vectors at all time points tested. Notably, all eyes treated with scAAV8-GFP displayed degeneration of the outer segment of PR cells from day 14 whereas retinal integrity was preserved in all eyes for ssAAV8-GFP. IHC of retinal cross-sections confirmed efficient transduction of the RPE cells and some PR cells. Intraocular inflammation was not observed during the study.

Conclusions : Subretinal delivery of both ssAAV2.8 and scAAV2.8 caused efficient transduction of RPE cells in the porcine retina. scAAV2.8-GFP showed faster onset of GFP expression, but also led to degeneration of the outer segments of the PR cells at the tested dose, which is a significant concern for translational application. In contrast, ssAAV2.8 did not cause any observable intraocular inflammation or retinal degeneration and may be the choice for further studies evaluating gene therapy for retinal diseases.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

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