Human iPSC-Derived Retinas Recapitulate the Fetal CRB1 CRB2 Complex Formation and Demonstrate that Photoreceptors and Müller Glia Are Targets of AAV5

Quinn, Peter M J; Buck, Thilo M.; Mulder, Aat A.; Ohonin, Charlotte; Alves, Celso; Vos, Rogier M.; Bialecka, Monika; Herwaarden, Tessa van; Dijk, Elon H. C. van; Talib, Mays; Freund, Christian; Mikkers, Harald; Hoeben, Rob C.; Goumans, Marie–José; Boon, Camiel J F; Koster, Abraham J.; Lopes, Susana M. Chuva de Sousa; Jost, Carolina R.; Wijnholds, Jan

doi:10.1016/j.stemcr.2019.03.002

Cited by 79 publications

(136 citation statements)

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“…AAV is a highly efficient method of delivering therapeutic genes to photoreceptor cells and several AAV based gene therapy trials for monogenic inherited retinal dystrophies are ongoing (for review see ). AAV2/5 has been shown to effectively transduce photoreceptors in a variety of species inter alia mice (Palfi et al, 2012), non-human primates (Boye et al, 2012) and human retinal explants (Wiley et al, 2018, Quinn et al, 2019.…”

Section: Discussionmentioning

confidence: 99%

“…Unlike previous models in 2D, these 3D structures contain photoreceptors with morphologically identifiable features; including, inner segments rich in mitochondria, rudimentary outer segments with connecting cilia, and synaptic pedicles in addition to bipolar, Müller glia, ganglion and amacrine cells, synaptic layers and an outer limiting membrane, arranged in retinal layers (reviewed in (Castro et al, 2019, Capowski et al, 2019. These advanced models have proven to have many translational research applications, including transplantation studies (Pearson et al, 2016, Gonzalez-Cordero et al, 2017, probing mechanisms of retinal development (Phillips et al, 2014), and specific retinal disease modelling and testing efficacy of potential therapies in human photoreceptor cells (Parfitt et al, 2016, Deng et al, 2018, Schwarz et al, 2017, Sharma et al, 2017, Quinn et al, 2019. To date, however, they have not been used to model and rescue photoreceptor cell death.…”

Section: Introductionmentioning

confidence: 99%

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Modelling and rescue of RP2 Retinitis Pigmentosa using iPSC Derived Retinal Organoids

Lane¹,

Jovanović²,

Shortall

et al. 2020

Preprint

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Mutations in RP2 lead to a severe form of X-linked retinitis pigmentosa (XLRP). RP2 functions as a GTPase activating protein (GAP) for the small GTPase ARL3, which is essential for cilia function and for photoreceptor development and maintenance. The mechanisms of RP2 associated retinal degeneration in humans are poorly understood, and genetically engineered animal models of RP2 XLRP present with differing retinal phenotypes and slow degeneration suggesting potential species differences. Here, we developed CRISPR gene edited isogenic RP2 knock-out (RP2 KO) induced pluripotent stem cells (iPSC) and RP2 patient derived iPSC to produce 3D retinal organoids as a human retinal disease model. The isogenic RP2 KO retinal organoids and two unrelated RP2 patient iPSC lines produced retinal organoids with a defined photoreceptor cell layer (ONL) containing rod and cone photoreceptors. Strikingly the RP2 KO and RP2 patient derived organoids showed a thinning of the ONL by 180 days (D180) of culture, which was associated with a spike in cell death in the ONL at D150 of differentiation.RNA sequencing confirmed induction of cell death pathways in the RP2 null organoids at this stage. Photoreceptor cell death and ONL thinning was attributed to cells undergoing terminal rod cell differentiation characterized by reduced RHO expression and fewer rhodopsin immunoreactive photoreceptors. Gene augmentation with a human RP2 transgene in an AAV2/5 vector efficiently transduced RP2 KO organoids and led to high levels of RP2 expression in both rods and cones. Importantly, the viral transduction significantly increased ONL thickness and restored rhodopsin expression, suggesting rescue of the RP2 degeneration phenotype. These data show that 3D retinal organoids can be used to model molecular defects associated with inherited retinal disease, photoreceptor cell death and also to test potential therapies targeted to prevent photoreceptor degeneration.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Modelling and rescue of RP2 Retinitis Pigmentosa using iPSC Derived Retinal Organoids

Lane¹,

Jovanović²,

Shortall

et al. 2020

Preprint

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show abstract

“…Alongside studies demonstrating that different eye cup‐like structures including 3D retinal organoids may provide essential and fundamental insight into developmental and regenerative processes , reports of the utility of these human tissues in eye disease modeling are now emerging (Table ). Most eye diseases involve the dysfunction and degeneration of the retina with blindness as the final outcome, as effective treatments do not currently exist.…”

Section: Retinal Tissue As Disease Modelmentioning

confidence: 99%

“…The works of Phillips et al and Quinn et al demonstrated how hiPSCs‐derived retinal tissue derived from patients could be applied to study genes involved in retinogenesis. In the first study, the authors modeled microphthalmia caused by mutations in the Visual System Homeobox 2 ( VXS2 ) gene by comparing optic vesicles generated from microphthalmia hiPSCs and healthy donor hiPSCs, thereby providing a paradigm for elucidating transcription factors and signaling pathways triggered by this early development associated gene.…”

Section: Retinal Tissue As Disease Modelmentioning

confidence: 99%

“…Mutations in the Crumbs homolog‐1 ( CRB1 ) gene are linked to retinal dystrophies that exhibit high‐phenotypic variability. In the comparative study by Quinn et al , using human retinal organoids derived from hiPSCs and human fetal retina, they compared the onset of CRB protein expression in both tissues showing that fetal CRB complex formation is replicated in hiPSC‐derived retina.…”

Section: Retinal Tissue As Disease Modelmentioning

confidence: 99%

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Deciphering retinal diseases through the generation of three dimensional stem cell-derived organoids: Concise Review

et al. 2019

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Three‐dimensional (3D) retinal organoids, in vitro tissue structures derived from self‐organizing cultures of differentiating human embryonic stem cells or induced pluripotent stem cells, could recapitulate some aspects of the cytoarchitectural structure and function of the retina in vivo. 3D retinal organoids display huge potential for the investigation of the pathogenesis of monogenic hereditary eye diseases that are related to the malfunction or degeneration of photoreceptors or retinal ganglion cells by providing an effective in vitro tool with multiple applications. In combination with recent genome editing tools, 3D retinal organoids could also represent a reliable and renewable source of transplantable cells for personalized therapies. In this review, we describe the recent advances in human pluripotent stem cells‐derived retinal organoids, determination of their histoarchitecture, complexity, and maturity. We also discuss their application as a means to decipher the pathogenesis of retinal diseases, as well as the main drawbacks and challenges. Stem Cells 2019;37:1496–1504

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Current perspectives in Leber congenital amaurosis type 8 mouse modeling

Nahar

Cho

2022

Developmental Dynamics

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Mutations in the CRB1 (Crumbs homolog 1) cause rare retinal diseases like retinitis pigmentosa type 12 (RP12) and Leber congenital amaurosis type 8 (LCA8). RP12 results in progressively worsening peripheral vision, whereas LCA8 causes severe visual impairment at birth or in early life. While several mouse models have been proposed for RP12, few replicate the full spectrum of human LCA8 pathology, such as disorganized retinal layering, abnormal retinal thickening, pigmentary defects, hyperreflective lesions, and severely attenuated electroretinogram responses at birth. Six models have been proposed utilizing the Cre-loxP system to delete candidate genes in specific retinal cell types and developmental stages. The model ablating Crb1 and its homolog Crb2 (using mRx-Cre) from the beginning of the eye development is the most complete as it shows blindness during the eye-opening stage, pigmentary defects in the RPE, ganglion cell layer heterotopia, disruption of retinal lamination, and acellular patches. LCA8 represents a unique type of retinal dystrophy among LCA subtypes, driven by dysfunctional retinal progenitor cells during eye development. In contrast, other LCA types and RP12 are caused by photoreceptor defects. Therefore, the most accurate LCA8-like mouse model must target both alleles of the Crb1 and Crb2 genes in the optic vesicle or earlier.

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Human iPSC-Derived Retinas Recapitulate the Fetal CRB1 CRB2 Complex Formation and Demonstrate that Photoreceptors and Müller Glia Are Targets of AAV5

Cited by 79 publications

References 50 publications

Modelling and rescue of RP2 Retinitis Pigmentosa using iPSC Derived Retinal Organoids

Modelling and rescue of RP2 Retinitis Pigmentosa using iPSC Derived Retinal Organoids

Deciphering retinal diseases through the generation of three dimensional stem cell-derived organoids: Concise Review

Current perspectives in Leber congenital amaurosis type 8 mouse modeling

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