Dr Paulina Ocieczek

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Genetic eye disorders are an important cause of visual impairment in children and adults, affecting approximately 1 in 1,000 people worldwide.

 They can affect any part of the eye and may present in isolation (non-syndromic) or as a syndrome in association with systemic involvement.

The United States Food and Drug Administration approval in 2017 of Spark Therapeutics’ Luxturna gene therapy for patients with biallelic 

-related retinal dystrophy opened the door for multiple new therapeutic strategies. Significant advancements in the field of inherited retinal diseases (IRDs) are making genetic testing essential for every patient, as emerging therapies may become an option for them in the near future.

IRDs are some of the most prevalent genetic eye disorders, affecting over 2 million people globally. They encompass a broad spectrum of progressive and stationary conditions and are the most frequent cause of sight loss in working age populations, causing a significant socioeconomic impact.

Mutations in over 300 genes responsible for IRDs have been identified. All Mendelian inheritance patterns are seen with IRDs.

Retinitis pigmentosa (RP) is the most prevalent type of IRD and affects 1 in 3,000 people worldwide.

 It is characterised by progressive rod photoreceptor degeneration, resulting in nyctalopia and progressive constriction of the visual field in a concentric pattern and, at later stages, loss of central vision due to the involvement of cones. It may be an isolated non-syndromic condition or part of a syndrome.

Usher syndrome is an autosomal recessive condition characterised by RP, auditory impairment and a variable level of vestibular dysfunction. It is the most common cause of deaf–blindness worldwide, with a prevalence of 4–17 per 100,000, accounting for about 18% of RP cases.

The triad of signs seen on retinal examination in Usher syndrome is mid-peripheral bone spicules, a waxy pale optic disc and arteriole attenuation.

Clinically, Usher syndrome is divided into three subtypes, mainly based on the severity of the congenital sensorineural hearing loss and the presence of vestibular dysfunction. Usher syndrome type 2 (USH2) is the most frequent form and is characterised by adolescent-onset RP, moderate-to-severe hearing loss from birth and intact vestibular function.

Ten causative genes have been identified to date. Usher genes encode proteins expressed in the inner ear and retina involved in actin-based intracellular trafficking, scaffolding, cell adhesion and signalling.

Six genes have been identified for Usher syndrome type 1 (USH1), with 

 accounting for over half of USH1 cases. Three gene loci have been described for USH2, with the 

 gene being responsible for over 80% of USH2 cases. So far, one gene, 

 is the most prevalent causative gene in Usher patients worldwide. It is also responsible for 23% of non-syndromic RP.

 is a large, 72-exon gene located on the long arm of chromosome 1q41; it codes the usherin transmembrane protein, present in the retina and cochlea where it plays roles in the intracellular transport of proteins within photoreceptors and in supporting the correct development of the stereocilia of inner ear hair cells, respectively.

 include a heterogeneous spectrum of missense, nonsense, indels frameshift, splice-site, deletion and duplication. Common pathogenic variants are found in exon 13; the most frequent is a single nucleotide deletion in exon 13, c.2299delG, p.(Glu767Serfs*21), which is associated with exon splicing producing a truncated usherin, and which causes USH2. A second common variant in exon 13 is a missense mutation, c.2276G>T, p.(Cys759Phe), which can cause a non-syndromic RP.

-associated retinopathy is challenging because of its broad spectrum of mutations and the large gene size. Although no treatment has been approved yet, numerous therapeutic approaches are under development for Usher-related RP. The retina is an excellent candidate for therapeutic application because of its accessibility and immune privilege status, while the progressive sight loss of RP makes it an important therapeutic target.

Approaches for treating RP in Usher syndrome

(A) Widefield colour imaging of the right fundus of a patient with 

related Usher syndrome showing bone spicules pigmentation in the mid-periphery, arteriole attenuation and optic disc pallor.

(B) Widefield autofluorescence imaging of the right fundus of a patient with 

-related Usher syndrome showing ring of the hyperautofluorescence in the macula.

Images courtesy of Dr Paulina Ocieczek and Prof. Mariya Moosajee)

Current therapeutic approaches include gene replacement; gene editing; nonsense suppression; antisense oligonucleotides; small-molecule therapies; and non-gene-specific therapies. Gene replacement using adeno-associated viral vectors (AAVs) has proven safe and effective in patients; however, its limited capacity of 4.7 kb means it is not suitable for carrying the 15.7 kb 

 cDNA. Dual hybrid AAV has a cargo capacity of 14 kb, which is still not sufficient, and its efficacy is limited.

 cDNA mediated by lentiviral-based equine infectious anaemia virus—UshStat—was the subject of a trial (NCT01505062) that was terminated early by its sponsor Sanofi following a review of clinical priorities. A second study evaluating the long-term safety of Ushstat is ongoing.

Work on non-viral DNA plasmid vectors is ongoing at the University College London Institute of Ophthalmology with the generation of robust plasmids that have successfully incorporated the entire 15.7 kb 

 cDNA and shown proof of-principle for production of usherin in zebrafish and human cell models. These vectors have a lower transfection efficacy compared with AAV, hence work is underway to couple the DNA plasmids with nanoparticles to improve penetration into photoreceptors.

Gene editing, which uses nuclease enzymes to cut around point mutations, small indels and splice site mutations, and recombination with template DNA containing a wild-type sequence to correct mutations in DNA, is suitable for any gene size. This approach is promising for large genes such as 

Clustered regularly interspaced short palindromic repeats (CRISPR/Cas), a gene-editing system, has been used for successful 

 editing of c.2299delG and c.2276G>T mutations in 

patient-derived fibroblasts and pluripotent stem cells. Eliminating off-target mutations and improving efficacy in post-mitotic photoreceptor cells remains a challenge for this approach.

 pathogenic variants are nonsense, and therefore could be an ideal target for translational readthrough-inducing drugs (TRIDs). These are small molecules such as ataluren or designer aminoglycosides (NB124) that bind to the translational machinery and insert amino acids at the site of premature stop codons, instead of release factors, thus restoring up to 25% of functional protein. The use of ataluren for 

 nonsense mutations in human embryonic kidney and patient-derived fibroblasts showed an increase in expression of usherin compared with control.

An important and promising modality for IRD therapy is antisense oligonucleotides (ASOs). Synthetic ASOs are single-stranded nucleotides approximately 15–30 bases long. They can be designed to bind to target pre-mRNA or mRNA of disease-related genes by base pairing and modulate target gene expression by the restoration of correct splicing. Gene size is not an obstacle in this approach but it is relatively mutation-dependent, and the half-life of these molecules means they require re-administration.

ASOs have been under investigation in several clinical trials. In February 2022, a Phase 2/3 trial (Illuminate) investigating the use of intravitreal sepofarsen (QR-110) for the treatment of Leber congenital amaurosis caused by a splicing defect in the 

 gene showed no benefit compared with a sham procedure.

In November 2021, Phase 1 of the Aurora trial, which assessed the safety and tolerability of QR-1123 for patients with autosomal dominant RP due to a P23H mutation in the 

 gene, was completed and met its key objectives.

The use of ASOs is also being investigated in 

-related RP. An ASO called QRX-411 targets the deep intronic mutation c.7595_2144A>G in intron 40 of 

 gene, causing the introduction of pseudoexon 40 due to incorrect splicing. Administration of QRX 411 was shown to partially restore correct splicing in patient-derived fibroblasts and in an optic cup model in preclinical work.

Most recently, a double-masked, randomised, controlled, multiple-dose Phase 2/3 clinical trial (Sirius; NCT05158296) of QR-421a (ultevursen) has been initiated at Moorfields Eye Hospital in London and several other centres worldwide. The aim is to target RP associated with 

 mutations in exon 13. Considering that the two most common 

mutations are located in exon 13, the success of this therapeutic approach would help a large number of syndromic and non-syndromic 

QR-421a is designed to facilitate in-frame skipping of exon 13 in the 

 gene during pre-mRNA splicing to restore a shorter but functional protein. The production of a shortened version of usherin without exon 13 has been demonstrated in patient-derived retinal organoids, with partial restoration of protein expression also observed in treated zebrafish larvae.

The Phase 1/2 Stellar study, which evaluated safety, tolerability and efficacy of a single injection of QR-421a in adult subjects, demonstrated benefit in treated versus control eyes in multiple visual parameters. The injection was well tolerated and no serious adverse events were reported.

Sirius aims to establish the safety and efficacy of multiple doses of QR-421a over 24 months in patients aged 12 and over with advanced vision loss. The trial aims to recruit 81 patients with vision worse then 20/40 and its primary endpoint is a change in best corrected visual acuity at 18 months compared with control.

There is a huge range of new therapeutic approaches and clinical trials for genetic eye diseases. Genetic testing of patients is now paramount because of the range of gene- and mutation-specific treatments that are emerging. These therapies hold much promise and are giving hope to a community of patients that previously had no therapeutic options available to them.

Dr Ocieczek completed her ophthalmology training in Warsaw, Poland, and is now a clinical research fellow at Moorfields Eye Hospital, London, UK. She has no conflicts of interest to declare.

Mariya Moosajee, MBBS, BSc, PhD, FRCOphth

Prof. Moosajee is a consultant ophthalmologist in genetic eye disease at Moorfields Eye Hospital and Great Ormond Street Hospital for Children, professor of molecular ophthalmology at UCL Institute of Ophthalmology and group leader of Ocular Genomics and Therapeutics at the Francis Crick Institute in London. She was named as one of the top 100 most influential people in ophthalmology in The Ophthalmologist Power List 2022. She is the site principal investigator for the ProQR Sirius Study at Moorfields Eye Hospital and has received honoraria from ProQR.

1. Méjécase C, Malka S, Guan Z, et al. Practical guide to genetic screening for inherited eye diseases. 

2. Toms M, Pagarkar W, Moosajee M. Usher syndrome: clinical features, molecular genetics and advancing therapeutics. 

3. ProQR Announces top-line results from phase 2/3 Illuminate trial of sepofarsen in CEP290-mediated LCA10. 

www.globenewswire.com/news-release/2022/02/11/2383604/33039/en/ProQR-Announces-Top-Line-Results-from-Phase-2-3-Illuminate-Trial-of-Sepofarsen-in-CEP290-mediated-LCA10.html

4. Positive results of QR-421a phase 1/2 clinical trial for Usher syndrome and non-syndromic retinitis pigmentosa. 

www.proqr.com/files/2021-03/Community%20Statement%20QR-421a%20program%20results%20March2021_0.pdf

5. Sirius - phase 2/3 clinical trial for USH2A mediated retinitis pigmentosa. 

www.proqr.com/sirius-and-celeste-studies-for-ush2a-retinitis-pigmentosa

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Increasing understanding of the genetics behind Usher syndrome suggests that treatment may become possible in the very near future.

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