Search Results (223)

Objectives: This study investigated a variant, RPGR NM_001034853.2 c.1307G>A, p.[Gly436Asp, p?], in a large Irish pedigree with severe X-Linked Retinitis Pigmentosa (XLRP). The effect of the variant on RNA splicing was interrogated using in vitro functional analysis to provide evidence of disease causality. Methods: Three related individuals presenting with XLRP underwent target-capture sequencing, together with confirmatory Sanger sequencing and cascade analyses, to identify candidate variants. In silico investigations were undertaken using SpliceAI (version 1.3.1) and Alamut Visual software (version 2.13), among others. Functional analyses using in vitro midigene splice assays employing gateway expression vectors were undertaken. Variant and wildtype RNA were amplified by RT-PCR to investigate effects on splicing. RPGR c.1307G>A was subsequently reclassified using ACMG/AMP and ClinGen SVI recommendations. Results: Midigene investigation confirmed a cryptic acceptor site is being utilised together with the cryptic branchpoint motif to excise intron 10 and 90 bases of exon 11, leading to a frameshift and the creation of a premature stop codon. No functional RPGR transcript is predicted to remain. Given evidence of aberrant splicing, the variant classification was upgraded to pathogenic. Conclusions: RPGR c.1307G>A leads to creation of a cryptic branchpoint within an exon, resulting in protein truncation with deleterious effect(s). To the best of our knowledge, this is the first variant that leads to creation of a cryptic branchpoint within an exon associated with any IRD. The results illustrate the importance of investigating the functional consequences of both coding and non-coding variants with a predicted impact on splicing to understand their pathogenicity. Full article

Mitochondrial dysfunction and oxidative stress are increasingly recognized as key contributors to the development and progression of retinal degenerative diseases, including age-related macular degeneration and inherited retinal dystrophies. Growing evidence suggests that alterations in mitochondrial function, excessive production of reactive oxygen species, defective mitophagy, and chronic inflammatory responses are closely interconnected processes that contribute to retinal cell damage and degeneration. This review provides an overview of the current understanding of the molecular mechanisms linking mitochondrial dysfunction to retinal degeneration, with particular emphasis on the impact of oxidative stress, mitochondrial quality-control pathways, and inflammatory signaling. Available evidence indicates that mitochondrial DNA damage, impaired bioenergetics, and dysregulated mitochondrial dynamics play a crucial role in the degeneration of photoreceptors and retinal pigment epithelium cells. In turn, oxidative stress further exacerbates mitochondrial impairment, creating a self-sustaining cycle that promotes disease progression. Recent advances have also highlighted the therapeutic potential of targeting mitochondrial pathways. Although several mitochondria-directed strategies have shown encouraging results in experimental models, their translation into clinical practice remains at an early stage. Overall, the available data identify mitochondria as a promising therapeutic target and support the development of precision medicine approaches aimed at preserving retinal function and slowing disease progression in patients with retinal degenerative disorders. Full article

Retinitis pigmentosa (RP) is a genetically heterogeneous group of inherited retinal degenerations with primary degeneration of rod photoreceptors followed by secondary cone loss. We investigated whether downregulating Nrl (neural retina leucine zipper), a key transcription factor specifying rod fate, can reprogram rods into a more resilient state. In a transgenic Nrl^N/N mouse in which Nrl was markedly downregulated, the rod phenotype became more like a rod precursor, particularly in the inferior retina. Crossing Nrl^N/N mice with two rod degeneration models, rd1 (Pde6b^rd1/rd1) and rhodopsin P23H knock-in (Rho^P23H/P23H) mice, showed significantly improved photoreceptor survival in double-mutant mice. In addition, AAV-mediated delivery of shRNA targeting Nrl mRNA substantially enhanced photoreceptor survival in rd10 (Pde6b^rd10/rd10) mice. These findings demonstrate that downregulation of Nrl reprograms rods and confers broad resistance to degeneration across multiple RP models. AAV-mediated Nrl knockdown represents a promising mutation-independent therapeutic strategy for autosomal recessive and dominant forms of RP. Full article

Inherited retinal dystrophies (IRDs) comprise a diverse group of genetic disorders that frequently result in irreversible vision loss due to photoreceptor dysfunction or degeneration. Among them, retinitis pigmentosa (RP) and achromatopsia (ACHM) are, in some cases, associated with pathogenic variants in PDE6A and PDE6C, respectively, which are key components of the phototransduction cascade. As most of IRDs still lack effective therapies, retinal organoids (ROs) provide a valuable in vitro model for the investigation of disease-associated mechanisms. Here, we generated induced pluripotent stem cell (iPSC)-derived ROs from an RP patient carrying compound heterozygous PDE6A mutations and from a patient with ACHM harboring a homozygous PDE6C mutation, along with their corresponding CRISPR/Cas9-corrected isogenic controls, which, to our knowledge, represent the first patient-derived RO models reported for the PDE6A and PDE6C genes. The mutant PDE6A line exhibited impaired neuroretinal vesicle formation and RO differentiation; however, a subset of RP-derived ROs matured appropriately and retained photoreceptor features. Moreover, the specific isoform expression pattern detected in retinal tissues reflected differences across developmental maturation stages that could influence disease severity. In contrast, the PDE6C_mutant ROs displayed normal structure and maturation, although cGMP hydrolysis within photoreceptors was likely compromised. In both models, CRISPR/Cas9-mediated correction restored the disease-associated phenotype resembling wild-type ROs. Collectively, these findings provide new insights into PDE6-associated pathogenesis, underscore the utility of patient-specific and gene-corrected ROs for elucidating IRD mechanisms, and support gene editing as a promising therapeutic strategy. Full article

Background: Non-syndromic retinitis pigmentosa (RP) is characterized by rod–cone degeneration, resulting in night blindness, visual field constriction, and eventual blindness. Recessively inherited RP is predominantly exacerbated in consanguineous populations, such as Pakistan. This study aimed to perform the genetic analysis of sixteen non-syndromic RP segregating Pakistani families, and to summarize the mutation spectrum of non-syndromic RP in our population by reviewing related literature. Methods: We screened 16 non-syndromic RP families using targeted capture panel sequencing of 344 genes related to inherited retinal dystrophies. Variants were prioritized based on rarity (minor allele frequency (MAF) < 0.001 in the gnomAD South Asian subset), pathogenicity assessments using ACMG/AMP criteria, and REVEL scores (>0.5). Candidate variants were validated for familial segregation through Sanger sequencing. Results: We identified 15 distinct variants across 14 genes associated with non-syndromic retinitis pigmentosa, comprising 6 missense, 7 nonsense, 1 frameshift, and 2 splice-site variants, including 4 novel variants, i.e., p.(Val220Met) and p.(Pro1282SerfsTer2) in RP1, 1 each in PDE6B (c.2021+5G>A), and PRCD p.(Ser38Ter). Homozygosity predominated, underscoring the impact of consanguinity on the burden of autosomal recessive disease in the present cohort, while the CERKL disease-causing mutation, i.e., p.(Arg257Ter), recurred in two families. Conclusions: This study expands Pakistan’s non-syndromic RP mutational spectrum by identifying novel variants in RP1, PDE6B, and PRCD, alongside recurrent CERKL and RHO mutations of the local population. The literature review suggests that RP1, TULP1, and PDE6B are among the most mutated genes in our population, supporting the value of population-specific genetic panels to enhance diagnostics and carrier screening. Full article

Prominin-1 (Prom1/CD133) has long been recognized as a structural determinant of photoreceptor outer segment (OS) morphogenesis, yet rapidly accumulating evidence extends its role to retinal pigment epithelium (RPE) homeostasis, encompassing autophagy–lysosomal flux, outer segment phagocytosis, mitochondrial function, and regulation of inflammatory stress. This review synthesizes mechanistic and transcriptomic insights that position PROM1 as a central regulator of photoreceptor and RPE integrity, reframing Prom1 disease as a multi-compartment retinal disorder relevant to both inherited retinal dystrophies (IRDs) and atrophic age-related macular degeneration (aAMD). We develop a dual-axis conceptual model in which Prom1 dysfunction can initiate pathology in either the photoreceptors (OS morphogenesis failure) or the RPE, including impaired autophagic flux, lysosomal activity, defective phagocytosis, and Epithelial-Mesenchymal Transition (EMT)-like de-differentiation, with secondary cross-compartmental degeneration. Clinically, autosomal-dominant missense variants associate with macular or cone-rod dystrophy, whereas biallelic truncating/splice-site mutations drive early-onset rod–cone disease and panretinal/RPE atrophy, illustrating genotype–phenotype diversity. By integrating recent high-resolution transcriptomic data from Prom1-deficient RPE cells with long-standing insights into photoreceptor biology, we highlight converging pathways of degeneration that challenge a photoreceptor-centric view and unify disparate phenotypes within a single molecular framework. These insights broaden the therapeutic landscape, advancing gene augmentation and pathway-targeted strategies to preserve RPE integrity, sustain photoreceptor function, and modify disease course in PROM1-associated IRDs and atrophic AMD. Full article

CRISPR–Cas9 has progressed from an experimental tool to a therapeutic modality, marked by the first regulatory approvals of an ex vivo-edited autologous CD34+ hematopoietic stem cell product that induces fetal hemoglobin (CASGEVY/exa-cel). In this narrative review, we synthesize modality-specific molecular diagnostic strategies used across early CRISPR clinical translation. In parallel, early clinical experience has begun to demonstrate the feasibility of in vivo editing, including subretinal delivery for CEP290-associated inherited retinal degeneration (EDIT-101 programme) and hepatocyte-targeted lipid nanoparticles (LNPs) for liver-derived targets such as transthyretin and plasma prekallikrein (KLKB1). As translation expands across hematologic, metabolic, ocular and oncology indications, development is increasingly constrained by the predictability and safety of editing outcomes, delivery-determined biodistribution and exposure time, and immune recognition of bacterial Cas9 orthologs and delivery components. We summarize diagnostic readouts for confirming patient genotype, quantifying on-target editing and expression changes, assessing off-target and structural outcomes using orthogonal assays, and monitoring clonal dynamics and immune responses during long-term follow-up. We also discuss how these readouts interface with CMC controls and regulatory expectations for advanced therapy medicinal products (ATMPs), highlighting the need for fit-for-purpose, standardized testing frameworks in early trials. Full article

Background: Retinal and optic nerve disorders are a leading cause of irreversible visual impairment worldwide. Advances in molecular genetics—including next-generation sequencing, genome-wide association studies, and gene-based therapeutic technologies—have reshaped understanding of both inherited and complex retinal diseases. However, translating genetic discovery into sustained clinical benefit remains biologically and practically constrained. Methods: A structured literature search was conducted using PubMed and Scopus to identify relevant studies published between 2015 and 2025. The search focused on molecular genetics, epigenetic modulation, mitochondrial biology, and translational applications in inherited retinal dystrophies and selected complex retinal diseases, prioritizing high-impact original research and systematic reviews addressing diagnostic innovation and therapeutic development. Results: Inherited retinal dystrophies represent the most advanced model of precision ophthalmology, with diagnostic yields approaching 70–80% in well-characterized cohorts. Gene augmentation and genome-editing strategies have demonstrated proof-of-concept efficacy, yet clinical benefit depends on residual cellular viability, delivery efficiency, and durability of expression. Emerging platforms include AAV-mediated gene transfer, in vivo CRISPR-based editing, RNA-directed splice modulation, and mitochondrial-targeted approaches. Persistent barriers include unresolved non-coding and structural variants, variant interpretation uncertainty, and endpoint selection in clinical trials. In contrast, complex retinal diseases such as glaucoma, age-related macular degeneration, and pathological myopia reflect polygenic susceptibility interacting with environmental and aging-related factors. Although polygenic risk scores refine probabilistic prediction, their utility is limited by ancestry bias and incomplete predictive performance. Epigenetic and mitochondrial mechanisms further modulate disease expression but remain largely non-actionable in routine practice. Conclusions: Retinal genetics has progressed from gene discovery to early therapeutic implementation. Future advances will depend on improved variant detection, functional validation, biomarker-guided staging, and integration of genomics with imaging and longitudinal modeling to achieve durable and equitable precision ophthalmology. Full article

Background/Objectives: Inherited retinal diseases (IRDs) represent a genetically heterogeneous group of disorders caused by mutations in over 280 genes with more than 3100 identified variants. While gene-specific replacement therapies have achieved landmark success with voretigene neparvovec (Luxturna) for biallelic RPE65-associated retinal dystrophy, developing individual therapies for each genetic subtype remains impractical. This review examines gene-agnostic therapeutic approaches utilizing neuroprotection and immunomodulation that target common pathophysiological mechanisms shared across multiple IRD genotypes. Methods: We reviewed the literature on neuroprotective and immunomodulatory gene therapy strategies for IRDs, focusing on neurotrophic factors and complement system modulation. Results: Neuroprotective approaches delivering neurotrophic factors—including pigment epithelium-derived factor (PEDF), ciliary neurotrophic factor (CNTF), rod-derived cone viability factor (RdCVF), brain-derived neurotrophic factor (BDNF), fibroblast growth factors (FGFs), glial cell line-derived neurotrophic factor (GDNF), and proinsulin—have demonstrated photoreceptor preservation across multiple preclinical IRD models regardless of the underlying genetic mutation. The recent FDA approval of CNTF cell-based gene therapy (Encelto) for macular telangiectasia type 2 validates this therapeutic paradigm. Complement system inhibition represents another gene-agnostic strategy, with intravitreal complement inhibitors approved for geographic atrophy secondary to age-related macular degeneration and gene therapy approaches targeting C3, C5, or delivering soluble complement regulators under investigation for IRDs. Combination strategies simultaneously addressing multiple pathogenic pathways may offer synergistic benefits. Conclusions: Gene-agnostic approaches targeting neuroprotection and immunomodulation offer a therapeutic paradigm capable of benefiting patients across the spectrum of IRD genotypes, potentially transforming treatment for conditions where mutation-specific therapies remain unavailable. Full article

Mitochondrial reactive oxygen species (mtROS) play a dual role in retinal physiology, acting as essential redox signalling mediators under homeostatic conditions but driving oxidative damage and neurodegeneration once regulatory thresholds are exceeded. Owing to the exceptionally high energetic demands of retinal neurons and supporting cells, even subtle perturbations in mitochondrial redox balance can precipitate progressive retinal dysfunction. Increasing evidence indicates that retinal neurodegenerative diseases, including glaucoma, diabetic retinopathy (DR), age-related macular degeneration (AMD), and inherited optic neuropathies, are characterised not by uniform oxidative stress, but by disease- and stage-specific mtROS signatures shaped by mitochondrial quality control capacity. This review synthesises current insights into the sources, regulation, and signalling functions of mtROS in the retina, with particular emphasis on threshold-dependent redox transitions, reverse electron transport, and the progressive failure of mitochondrial quality control mechanisms, including mitophagy, mitochondrial dynamics, and redox-responsive transcriptional networks. The limitations of non-selective antioxidant strategies are critically examined, highlighting why indiscriminate ROS suppression has yielded limited clinical benefit. In contrast, emerging therapeutic approaches aimed at recalibrating mitochondrial redox homeostasis, rather than abolishing physiological signalling, are discussed in the context of disease stage, metabolic state, and mitochondrial competence. By integrating redox biology with mitochondrial quality control and precision medicine concepts, this review proposes a unifying framework in which retinal neurodegeneration is governed by regulated mtROS signalling and the progressive exhaustion of mitochondrial resilience. This model defines critical therapeutic windows for mitochondria-targeted intervention and provides a framework for biomarker-guided patient stratification. Full article

Genome editing has emerged as a transformative approach for understanding and treating retinal degenerative diseases. Combining this technology with pluripotent stem cells provides an ideal platform for modeling human development and disease, and investigating emerging therapeutic strategies ultimately aimed towards in vivo correction. This approach enables both functional studies to understand retinal degeneration and the early development of targeted therapies for inherited disease. This review offers a comprehensive overview of genome-editing techniques and the ability to create new clinically relevant models to understand human disease in retinal research, focusing on the use of the CRISPR-Cas9 system in induced pluripotent stem cells (iPSCs) and embryonic stem cells (ESCs), as well as highlighting recent advancements in base and prime editing. Gene editing in various retinal diseases is discussed in context of studies focusing on disease modeling or developing therapeutic strategies. Continued refinement of these techniques will be essential for advancing translational applications in retinal disease treatment. Full article

Retinitis pigmentosa (RP) comprises a heterogeneous group of inherited retinal dystrophies characterized by the progressive degeneration of photoreceptors, leading to night blindness and gradual loss of peripheral vision. RP is characterized by a substantial genetic heterogeneity, with more than 85 genes implicated across autosomal dominant, autosomal recessive, and X-linked inheritance patterns. Recent studies have identified mutations in the ARL3 gene as a causative factor in both syndromic and non-syndromic forms of RP, including autosomal dominant and recessive cases. ARL3 encodes a small GTPase that plays a crucial role in intracellular trafficking, particularly within photoreceptors. This process is critical for maintaining ciliary function and phototransduction. Here, we investigate the pathogenic mechanisms of the ARL3 c.199G>C (p.Asp67His) variant identified in individuals from a four-generation family. We show that mutant ARL3 disrupts normal protein expression and affects ciliogenesis. Clinically affected individuals showed a non-syndromic retinal degenerative RP phenotype, with marked intrafamilial heterogeneity, ranging from extensive retinal atrophy to the absence of clinical manifestation, independent of age. This report highlights the incomplete penetrance and variable expressivity associated with the ARL3 variant and emphasizes the value of combining molecular diagnostics with functional validation to expedite molecular diagnosis. Full article

Inherited retinal degenerations, such as retinitis pigmentosa, are a leading cause of irreversible vision loss, yet broadly effective treatments remain elusive. Impaired cellular waste clearance via autophagy–lysosomal pathways have been implicated in photoreceptor death, but the spatiotemporal dynamics of these processes during degeneration remain poorly understood. Using the rd10 mouse model of retinitis pigmentosa, we characterised autophagy–lysosomal dysfunction at key stages of photoreceptor degeneration (postnatal day P17, P22, P35) through super-resolution imaging of RFP-EGFP-LC3 reporter mice, Western blot, and bulk RNA sequencing. Autophagosome and autolysosome numbers were significantly elevated across all photoreceptor compartments (inner/outer segments, outer nuclear layer, outer plexiform layer) at P17, prior to significant photoreceptor nuclei loss. Autophagosome and autolysosome size progressively increased from P22 onwards, suggesting accumulation of unprocessed intracellular waste. Molecular analyses revealed downregulation of mTOR protein, upregulation of autophagy-related genes, and increased lysosomal processes from P17. These histological and molecular findings are consistent with early autophagy induction followed by overwhelmed degradative capacity. Our findings identify autophagy–lysosomal change as an early event in photoreceptor loss in the rd10 model, revealing a critical therapeutic window for mutation-independent interventions targeting cellular clearance pathways in inherited retinal degenerations. Full article

Ophthalmic diseases, including inherited retinal dystrophies, age-related macular degeneration (AMD), and glaucomatous neuropathies, are often driven by the expression of pathogenic proteins or dysfunctional non-coding RNAs that are currently considered ‘undruggable’ with conventional small-molecule therapeutics. The emerging strategy of Ribonuclease-Targeting Chimeras (RIBOTACs) offers a revolutionary approach to address this therapeutic gap. RIBOTACs are heterobifunctional small molecules designed to bind a specific target RNA with one moiety and recruit a latent endogenous ribonuclease, such as RNase L, with the other, thereby catalyzing the RNA’s degradation. This targeted degradation can potentially halt the production of mutant proteins, eliminate toxic gain-of-function RNAs, or modulate key regulatory pathways involved in angiogenesis, inflammation, and apoptosis—core processes in many blinding diseases. This review explores the immense potential of applying RIBOTAC technology to ophthalmology, discussing prospective targets such as mutant alleles in retinitis pigmentosa, VEGF transcripts in neovascular AMD, and inflammatory mediators in uveitis. We will also address the unique challenges and opportunities for RIBOTAC development in the eye, including delivery strategies to overcome ocular barriers, the need for high specificity to avoid off-target RNA degradation, and the optimization of pharmacokinetic properties for intraocular administration. With continued innovation, RIBOTACs are poised to evolve into a robust therapeutic platform, expanding the druggable genome and enabling precise, durable treatments for a range of currently intractable ophthalmic conditions. Full article

Background/Objectives: Retinitis pigmentosa is a degenerative retinal disease and a major cause of inherited blindness globally. The pro-survival kinase AKT is downregulated in degenerating photoreceptors in retinitis pigmentosa, and its activation has shown neuroprotective effects in retinitis pigmentosa and other neurodegenerative disorders. In this study, we evaluated the therapeutic potential of SC79, a pharmaceutical AKT activator, in two mouse models of retinitis pigmentosa, rd1.GFP and RhoP23H.GFP. Methods: SC79 was administered intravitreally at postnatal day 12 (P12) and analysis was conducted at P16. Results: SC79 at 10 µM was well tolerated in wildtype mice, with no reduction in retinal function or thickness. In rd1.GFP mice, SC79 partially preserved peripheral outer nuclear layer (ONL) thickness, improved rod photoreceptor-driven optomotor contrast sensitivity responses, and improved cone photoreceptor morphology. Immunohistochemistry of retinal sections indicated AKT-related protein expression changes in both sham and SC79-treated rd1.GFP retinas, with sham injections leading to decreases in this pathway and SC79 injections restoring this back to uninjected protein levels or higher, indicating the damage from intravitreal injections can induce AKT-related protein expression changes. In RhoP23H.GFP mice, changes to the visual response from the therapeutic effects of SC79 were not detectable. An increased dosage of SC79 at 100 µM was evaluated in wildtype mice and showed no major toxic effects, although it did not confer neuroprotective benefits in either disease model. Conclusions: These results demonstrate the potential therapeutic effect of AKT pathway modulation for preserving photoreceptors in recessive retinitis pigmentosa, with further optimisation of treatment delivery required. Full article