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Cited by 36Virus-based gene therapy researchRetinal Development and DisordersRNA Interference and Gene Delivery

Tropism of AAV Vectors in Photoreceptor-Like Cells of Human iPSC-Derived Retinal Organoids

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McClements, Michelle E., Steward, Hannah, Atkin, William, Goode, Emily Archer, Gándara, Carolina, Chichagova, Valeria, MacLaren, Robert E.·2022·Translational Vision Science & Technology
DOI 10.1167/tvst.11.4.3PMC PMC8994202
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Abstract

Purpose: To expand the use of human retinal organoids from induced pluripotent stem cells (iPSCs) as an in vitro model of the retina for assessing gene therapy treatments, it is essential to establish efficient transduction. To date, targeted transduction of the photoreceptor-like cells of retinal organoids with adeno-associated virus (AAV) vectors has had varied degrees of success, which we have looked to improve in this study. Methods: Retinal organoids were differentiated from iPSCs of healthy donors and transduced with reporter AAV containing a CAG.GFP, CAG.RFP, GRK1.GFP, or EFS.GFP transgene. Capsid variants assessed were AAV5, AAV2 7m8, AAV2 quad mutant, AAV2 Y444F, and AAV8 Y733F. At 27 days post-transduction, retinal organoids were assessed for reporter expression and viability. Results: The short intron-less elongation factor 1 alpha (EFS) promoter provided minimal reporter expression, whereas vectors containing the CAG promoter enabled transduction in 1% to 37% of cells depending on the AAV serotype; the AAV2 quad mutant (average 19.4%) and AAV2 7m8 (16.4%) outperformed AAV5 (12%) and AAV8 Y733F (2.1%). Reporter expression from rhodopsin kinase (GRK1) promoter transgenes occurred in ∼5% of cells regardless of the serotype. Positive co-localization with recoverin-expressing cells was achieved from all GRK1 vectors and the CAG AAV2 quad mutant variant. Treatment with the AAV vectors did not influence retinal organoid viability. Conclusions: Reliable transduction of the photoreceptor-like cells of retinal organoids can be readily achieved. When using a CAG-driven transgene, transduction of a broad range of cell types is observed, and GRK1 transgenes provide a more restricted expression profile locating to the outer layer of photoreceptor-like cells of retinal organoids. Translational Relevance: This study expands the AAV capsid and transgene options for preclinical testing of gene therapy in iPSC-derived human retinal organoids.

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  • AAV2.7m8 is a useful delivery vector for cells across retinal cell types.

    “Overall, our results indicated AAV2.7m8 to be a useful delivery vector for cells across retinal cell types, confirming several previous studies (29–32).”

    From Characterization of RNA editing and gene therapy with a compact CRISPR-Cas13 in the retina (2024) · cites this paper as [31]

    The Results (Extent of Vector Transduction in Retinal Organoids; page not provided) describe broad AAV2.7m8 transduction in human retinal organoids. The Discussion calls its transduction robust with the CAG promoter, but reports minimal expression with EFS. Thus the source supports its usefulness in retinal organoids under tested conditions, not an unqualified claim of delivery across retinal cell types. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • AAV2.7m8 is effective as a vector for retinal transduction and supports its clinical utility.

    “confirming its effectiveness as a vector for retinal transduction and supporting its clinical utility (28, 31).”

    From Characterization of RNA editing and gene therapy with a compact CRISPR-Cas13 in the retina (2024) · cites this paper as [31]

    The Discussion reports robust AAV2 7m8 transduction in retinal organoids with the CAG promoter, and notes that the vector is being tested for safety and efficacy in humans. These preclinical results support its potential clinical use, but the paper does not establish clinical utility or effectiveness in patients; it also reports minimal expression with the EFS promoter. The indexed passages provide no page numbers. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • Due to their tissue-specific tropisms, some of the most common AAV serotypes in use for retinal gene therapy include AAV2, AAV4, AAV5, and AAV8.

    “Due to their tissue-specific tropisms, some of the most common AAV serotypes in use for retinal gene therapy include AAV2, AAV4, AAV5, and AAV8, as mentioned previously.26,110”

    From Gene Therapy for Retinal Degenerative Diseases: Progress, Challenges, and Future Directions (2023) · cites this paper as [110]

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Research works printed but not linked: 32

  1. [1]Cehajic-Kapetanovic J, Xue K, Martinez-Fernandez de la Camara C, et al. Initial results from a first-in-human gene therapy trial on X-linked retinitis pigmentosa caused by mutations in RPGR. Nat Med. 2020; 26: 354– 359. 32094925 PMC7104347
  2. [2]Xue K, MacLaren RE. Ocular gene therapy for choroideremia: clinical trials and future perspectives. Expert Rev Ophthalmol. 2018; 13: 129– 138. 31105764 PMC6520227
  3. [3]Dreismann AK, McClements ME, Barnard AR, et al. Functional expression of complement factor I following AAV-mediated gene delivery in the retina of mice and human cells. Gene Ther. 2021; 28: 265– 276. 33750925 PMC8149295
  4. [4]van Wyk M, Hulliger EC, Girod L, Ebneter A, Kleinlogel S. Present molecular limitations of ON-bipolar cell targeted gene therapy. Front Neurosci. 2017; 11: 11372. PMC5372788 28424574
  5. [6]Orlans HO, Edwards TL, Silva SRD, Patrício MI, MacLaren RE. Human retinal explant culture for ex vivo validation of AAV gene therapy. Methods Mol Biol. 2018; 1715; 289– 303. 29188522
  6. [7]De Silva SR, Issa PC, Singh MS, et al. Single residue AAV capsid mutation improves transduction of photoreceptors in the Abca4–/– mouse and bipolar cells in the rd1 mouse and human retina ex vivo. Gene Ther. 2016; 23: 767– 774. 27416076 PMC5097463
  7. [8]Eiraku M, Takata N, Ishibashi H, et al. Self-organizing optic-cup morphogenesis in three-dimensional culture. Nature. 2011; 472: 51– 56. 21475194
  8. [9]Zhong X, Gutierrez C, Xue T, et al. Generation of three-dimensional retinal tissue with functional photoreceptors from human iPSCs. Nat Commun. 2014; 5: 4047. 24915161 PMC4370190
  9. [10]Reichman S, Slembrouck A, Gagliardi G, et al. Generation of storable retinal organoids and retinal pigmented epithelium from adherent human iPS cells in xeno-free and feeder-free conditions. Stem Cells. 2017; 35: 1176– 1188. 28220575
  10. [11]Chichagova V, Dorgau B, Felemban M, et al. Differentiation of retinal organoids from human pluripotent stem cells. Curr Protoc Stem Cell Biol. 2019; 50: e95. 31479596
  11. [12]Chichagova V, Hilgen G, Ghareeb A, et al. Human iPSC differentiation to retinal organoids in response to IGF1 and BMP4 activation is line- and method-dependent. Stem Cells. 2020; 38: 195– 201. 31721366 PMC7383896
  12. [14]Gonzalez-Cordero A, Goh D, Kruczek K, et al. Assessment of AAV vector tropisms for mouse and human pluripotent stem cell-derived RPE and photoreceptor cells. Hum Gene Ther. 2018; 29: 1124– 1139. 29580100
  13. [15]Garita-Hernandez M, Routet F, Guibbal L, et al. AAV-mediated gene delivery to 3D retinal organoids derived from human induced pluripotent stem cells. Int J Mol Sci. 2020; 21: 994. PMC7036814 32028585
  14. [16]Völkner M, Pavlou M, Büning H, Michalakis S, Karl MO. Optimized adeno-associated virus vectors for efficient transduction of human retinal organoids. Hum Gene Ther. 2021; 32: 694– 706. 33752467
  15. [18]Lane A, Jovanovic K, Shortall C, et al. Modeling and rescue of RP2 retinitis pigmentosa using iPSC-derived retinal organoids. Stem Cell Reports. 2020; 15: 67– 79. 32531192 PMC7363745
  16. [19]Kallman A, Capowski EE, Wang J, et al. Investigating cone photoreceptor development using patient-derived NRL null retinal organoids. Commun Biology. 2020; 3: 82. PMC7035245 32081919
  17. [20]Gao M-L, Lei X-L, Han F, et al. Patient-specific retinal organoids recapitulate disease features of late-onset retinitis pigmentosa. Front Cell Dev Biol. 2020; 8: 128. 32211407 PMC7068133
  18. [21]Deng W-L, Gao M-L, Lei X-L, et al. Gene correction reverses ciliopathy and photoreceptor loss in iPSC-derived retinal organoids from retinitis pigmentosa patients. Stem Cell Reports. 2018; 10: 1267– 1281. 29526738 PMC5998840
  19. [22]Huang K-C, Wang M-L, Chen S-J, et al. Morphological and molecular defects in human three-dimensional retinal organoid model of X-linked juvenile retinoschisis. Stem Cell Reports. 2019; 13: 906– 923. 31668851 PMC6895767
  20. [23]Buskin A, Zhu L, Chichagova V, et al. Disrupted alternative splicing for genes implicated in splicing and ciliogenesis causes PRPF31 retinitis pigmentosa. Nat Commun. 2018; 9: 4234. 30315276 PMC6185938
  21. [24]Kruczek K, Qu Z, Gentry J, et al. Gene therapy of dominant CRX-Leber congenital amaurosis using patient stem cell-derived retinal organoids. Stem Cell Reports. 2021; 16: 252– 263. 33513359 PMC7878833
  22. [25]Cuevas E, Holder DL, Alshehri AH, Tréguier J, Lakowski J, Sowden JC. NRL−/− gene edited human embryonic stem cells generate rod-deficient retinal organoids enriched in S-cone-like photoreceptors. Stem Cells. 2021; 39: 414– 428. 33400844 PMC8438615
  23. [26]Kantor A, McClements ME, MacLaren RE. CRISPR-Cas9 DNA base-editing and prime-editing. Int J Mol Sci. 2020; 21: 6240. PMC7503568 32872311
  24. [27]Fry LE, McClements ME, MacLaren RE. Analysis of pathogenic variants correctable with CRISPR base editing among patients with recessive inherited retinal degeneration. JAMA Ophthalmol. 2021; 139: 319– 328. 33507217 PMC7844696
  25. [28]Fry LE, Peddle CF, Barnard AR, McClements ME, MacLaren RE. RNA editing as a therapeutic approach for retinal gene therapy requiring long coding sequences. Int J Mol Sci. 2020; 21: 777. PMC7037314 31991730
  26. [29]Peddle CF, Fry LE, McClements ME, MacLaren RE. CRISPR interference–potential application in retinal disease. Int J Mol Sci. 2020; 21: 2329. PMC7177328 32230903
  27. [32]Hallam D, Hilgen G, Dorgau B, et al. Human-induced pluripotent stem cells generate light responsive retinal organoids with variable and nutrient-dependent efficiency. Stem Cells. 2018; 36: 1535– 1551. 30004612 PMC6392112
  28. [33]Fripont S, Marneffe C, Marino M, Rincon MY, Holt MG. Production, purification, and quality control for adeno-associated virus-based vectors. J Vis Exp. 2019; 143: e58960. 30774140
  29. [34]Kantor A, McClements ME, Peddle CF, et al. CRISPR genome engineering for retinal diseases. Prog Mol Biol Transl Sci. 2021; 182: 29– 79. 34175046
  30. [36]Gaub BM, Berry MH, Holt AE, Isacoff EY, Flannery JG. Optogenetic vision restoration using rhodopsin for enhanced sensitivity. Mol Ther. 2015; 23: 1562– 1571. 26137852 PMC4817926
  31. [37]McClements ME, Staurenghi F, Visel M, et al. AAV induced expression of human rod and cone opsin in bipolar cells of a mouse model of retinal degeneration. BioMed Res Int. 2021; 2021: 1– 8. PMC7612646 35465048
  32. [40]Zhong L, Li B, Mah CS, et al. Next generation of adeno-associated virus 2 vectors: point mutations in tyrosines lead to high-efficiency transduction at lower doses. Proc Natl Acad Sci USA. 2008; 105: 7827– 7832. 18511559 PMC2402387

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Authors
McClements, M. E., Steward, H., Atkin, W., Goode, E. A., Gándara, C., Chichagova, V., & MacLaren, R. E.
Affiliations
  • University of Oxford (GB)
  • Newcells Biotech Limited, Newcastle upon Tyne, UK
Corresponding author
MacLaren, Robert E.
ORCID
Venue
Translational Vision Science & Technology
Publisher
Association for Research in Vision and Ophthalmology (ARVO)
Published
2022-04-04
Volume
11
Issue
4
Pages
3
ISSN
2164-2591 (ISSN-L)
DOI
10.1167/tvst.11.4.3
PMC
PMC8994202
OpenAlex
W4223627425
Cited by
36
References
8
Citations by year
2026: 10 · 2025: 7 · 2024: 9 · 2023: 9 · 2022: 1
FWCI
3.92
Type
journal article
Language
en
Open access
Licence
Locations
  • Translational Vision Science & Technology (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
National Institute for Health and Care Research (NIHR201338)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Retinal Development and Disorders
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
Keywords
adeno-associated virus vectors, retinal organoids, induced pluripotent stem cells, gene therapy, transduction efficiency, AAV5, CAG promoter, recoverin
MeSH terms
Dependovirus, Genetic Vectors, Humans, Organoids, Retina, Transduction, Genetic, Tropism, Induced Pluripotent Stem Cells

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