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Germline knockout of Nr2e3 protects photoreceptors in three distinct mouse models of retinal degeneration.

ABSTRACT: Retinitis pigmentosa (RP) is a common form of retinal dystrophy that can be caused by mutations in any one of dozens of rod photoreceptor genes. The genetic heterogeneity of RP represents a significant challenge for the development of effective therapies. Here, we present evidence for a potential gene-independent therapeutic strategy based on targeting Nr2e3, a transcription factor required for the normal differentiation of rod photoreceptors. Nr2e3 knockout results in hybrid rod photoreceptors that express the full complement of rod genes, but also a subset of cone genes. We show that germline deletion of Nr2e3 potently protects rods in three mechanistically diverse mouse models of retinal degeneration caused by bright-light exposure (light damage), structural deficiency (rhodopsin-deficient Rho^-/- mice), or abnormal phototransduction (phosphodiesterase-deficient rd10 mice). Nr2e3 knockout confers strong neuroprotective effects on rods without adverse effects on their gene expression, structure, or function. Furthermore, in all three degeneration models, prolongation of rod survival by Nr2e3 knockout leads to lasting preservation of cone morphology and function. These findings raise the possibility that upregulation of one or more cone genes in Nr2e3-deficient rods may be responsible for the neuroprotective effects we observe.

SUBMITTER: Kolesnikov AV

PROVIDER: S-EPMC10945761 | biostudies-literature | 2024 Mar

REPOSITORIES: biostudies-literature

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Germline knockout of Nr2e3 protects photoreceptors in three distinct mouse models of retinal degeneration.

Kolesnikov Alexander V AV Murphy Daniel P DP Corbo Joseph C JC Kefalov Vladimir J VJ

Proceedings of the National Academy of Sciences of the United States of America 20240305 11

Retinitis pigmentosa (RP) is a common form of retinal dystrophy that can be caused by mutations in any one of dozens of rod photoreceptor genes. The genetic heterogeneity of RP represents a significant challenge for the development of effective therapies. Here, we present evidence for a potential gene-independent therapeutic strategy based on targeting Nr2e3, a transcription factor required for the normal differentiation of rod photoreceptors. Nr2e3 knockout results in hybrid rod p ...[more]

PMID: 38442152

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Project description:BackgroundThe retinal degenerative diseases retinitis pigmentosa (RP) and atrophic age- related macular degeneration (AMD) are characterized by vision loss from photoreceptor (PR) degeneration. Unfortunately, current treatments for these diseases are limited at best. Genetic and other preclinical evidence suggest a relationship between retinal degeneration and inflammation. To further explore this relationship, we tested whether Ibuprofen (IBU), an FDA-approved non-steroidal anti-inflammatory drug (NSAID), could promote PR survival and function in a mouse model of light damage (LD)-induced PR degeneration.MethodsLD was induced by exposing mice to 4000 lx of light for 2-4 hours (h). IBU (100 or 200 mg/kg) or vehicle was administered by daily intraperitoneal injection. Retinal structure and function were evaluated by spectral-domain optical coherence tomography (SD-OCT) and electroretinography (ERG). Cell death genes were analyzed at 24 and 72 h after LD using the Mouse Pan-Cell Death Pathway PCR Array (88 genes). The cellular location and protein expression of key necroptosis genes were assessed by immunohistochemistry.ResultsRetinal outer nuclear layer (ONL) thickness in vehicle-injected LD animals was 8.7 ± 0.6% of retinas without LD (p < 0.0001). In IBU 200 mg/kg treated mice, central ONL thickness was 74.9 ± 7.7% of untreated retinas (p < 0.001). A-wave and b-wave ERG amplitudes were significantly preserved in IBU-treated animals. IBU significantly inhibited retinal inflammation. Twenty-four hour after LD, retinal mRNA expression for the inflammatory-factors tumor necrosis factor (Tnf), interleukin-1 beta (Il1B), and C-C motif chemokine ligand 2 (Ccl2) increased by 10-, 17-, and 533-fold, respectively; in IBU-treated animals, the expression levels of these inflammatory factors were not significantly different from no-LD controls. Expression of key necroptosis genes, including Ripk3 and Mlkl, were upregulated in LD vehicle-treated mice, but dramatically reduced to near no LD levels in LD IBU-treated mice. Microglia activation and MLKL protein upregulation were observed primarily in photoreceptors 12 h after LD, as assessed by immunohistochemistry. IBU reduced the upregulation of MLKL protein and microglia migration in the ONL and outer plexiform layer (OPL) of treated retinas.ConclusionsSystemic administration of the anti-inflammatory drug IBU partially protected mouse retinas from light-induced photochemical damage and inhibited both inflammation and the necroptosis cell death pathways. Our results suggest that NSAIDs may provide a promising therapeutic approach for treatment of the human retinal degenerative diseases.

Project description:BackgroundCEP290-associated inherited retinal degeneration causes severe early-onset vision loss due to pathogenic variants in CEP290. EDIT-101 is a clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) gene-editing complex designed to treat inherited retinal degeneration caused by a specific damaging variant in intron 26 of CEP290 (IVS26 variant).MethodsWe performed a phase 1-2, open-label, single-ascending-dose study in which persons 3 years of age or older with CEP290-associated inherited retinal degeneration caused by a homozygous or compound heterozygous IVS26 variant received a subretinal injection of EDIT-101 in the worse (study) eye. The primary outcome was safety, which included adverse events and dose-limiting toxic effects. Key secondary efficacy outcomes were the change from baseline in the best corrected visual acuity, the retinal sensitivity detected with the use of full-field stimulus testing (FST), the score on the Ora-Visual Navigation Challenge mobility test, and the vision-related quality-of-life score on the National Eye Institute Visual Function Questionnaire-25 (in adults) or the Children's Visual Function Questionnaire (in children).ResultsEDIT-101 was injected in 12 adults 17 to 63 years of age (median, 37 years) at a low dose (in 2 participants), an intermediate dose (in 5), or a high dose (in 5) and in 2 children 9 and 14 years of age at the intermediate dose. At baseline, the median best corrected visual acuity in the study eye was 2.4 log10 of the minimum angle of resolution (range, 3.9 to 0.6). No serious adverse events related to the treatment or procedure and no dose-limiting toxic effects were recorded. Six participants had a meaningful improvement from baseline in cone-mediated vision as assessed with the use of FST, of whom 5 had improvement in at least one other key secondary outcome. Nine participants (64%) had a meaningful improvement from baseline in the best corrected visual acuity, the sensitivity to red light as measured with FST, or the score on the mobility test. Six participants had a meaningful improvement from baseline in the vision-related quality-of-life score.ConclusionsThe safety profile and improvements in photoreceptor function after EDIT-101 treatment in this small phase 1-2 study support further research of in vivo CRISPR-Cas9 gene editing to treat inherited retinal degenerations due to the IVS26 variant of CEP290 and other genetic causes. (Funded by Editas Medicine and others; BRILLIANCE ClinicalTrials.gov number, NCT03872479.).

Project description:The visual cycle refers to a series of biochemical reactions of retinoids in ocular tissues and supports the vision in vertebrates. The visual cycle regenerates visual pigments chromophore, 11-cis-retinal, and eliminates its toxic byproducts from the retina, supporting visual function and retinal neuron survival. Unfortunately, during the visual cycle, when 11-cis-retinal is being regenerated in the retina, toxic byproducts, such as all-trans-retinal and bis-retinoid is N-retinylidene-N-retinylethanolamine (A2E), are produced, which are proposed to contribute to the pathogenesis of the dry form of age-related macular degeneration (AMD). The primary biochemical defect in Stargardt disease (STGD1) is the accelerated synthesis of cytotoxic lipofuscin bisretinoids, such as A2E, in the retinal pigment epithelium (RPE) due to mutations in the ABCA4 gene. To prevent all-trans-retinal-and bisretinoid-mediated retinal degeneration, slowing down the retinoid flow by modulating the visual cycle with a small molecule has been proposed as a therapeutic strategy. The present study describes RPE65-61, a novel, non-retinoid compound, as an inhibitor of RPE65 (a key enzyme in the visual cycle), intended to modulate the excessive activity of the visual cycle to protect the retina from harm degenerative diseases. Our data demonstrated that (±)-RPE65-61 selectively inhibited retinoid isomerase activity of RPE65, with an IC50 of 80 nM. Furthermore, (±)-RPE65-61 inhibited RPE65 via an uncompetitive mechanism. Systemic administration of (±)-RPE65-61 in mice resulted in slower chromophore regeneration after light bleach, confirming in vivo target engagement and visual cycle modulation. Concomitant protection of the mouse retina from high-intensity light damage was also observed. Furthermore, RPE65-61 down-regulated the cyclic GMP-AMP synthase stimulator of interferon genes (cGAS-STING) pathway, decreased the inflammatory factor, and attenuated retinal apoptosis caused by light-induced retinal damage (LIRD), which led to the preservation of the retinal function. Taken together, (±)-RPE65-61 is a potent visual cycle modulator that may provide a neuroprotective therapeutic benefit for patients with STGD and AMD.

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Germline knockout of Nr2e3 protects photoreceptors in three distinct mouse models of retinal degeneration.

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Germline knockout of <i>Nr2e3</i> protects photoreceptors in three distinct mouse models of retinal degeneration.

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