Extending diagnostic practices in gyrate atrophy: Enzymatic characterization and the development of an in vitro pyridoxine responsiveness assay.

Gyrate atrophy of the choroid and retina (GACR) is caused by pathogenic biallelic variants in the gene encoding ornithine-δ-aminotransferase (OAT), and is characterized by progressive vision loss leading to blindness. OAT is a pyridoxal-5'-phosphate (PLP) dependent enzyme that is mainly involved in ornithine catabolism, and patients with a deficiency develop profound hyperornithinemia. Therapy is aimed at lowering ornithine levels through dietary arginine restriction and, in some cases, through enhancement of OAT activity via supraphysiological dosages of pyridoxine. In this study, we aimed to extend diagnostic practices in GACR by extensively characterizing the consequences of pathogenic variants on the enzymatic function of OAT, both at the level of the enzyme itself as well as the flux through the ornithine degradative pathway. In addition, we developed an in vitro pyridoxine responsiveness assay. We identified 14 different pathogenic variants, of which one variant was present in all patients of Dutch ancestry (p.(Gly353Asp)). In most patients the enzymatic activity of OAT as well as the rate of [14C]-ornithine flux was below the limit of quantification (LOQ). Apart from our positive control, only one patient cell line showed responsiveness to pyridoxine in vitro, which is in line with the reported in vivo pyridoxine responsiveness in this patient. None of the patients harboring the p.(Gly353Asp) substitution were responsive to pyridoxine in vivo or in vitro. In silico analysis and small-scale expression experiments showed that this variant causes a folding defect, leading to increased aggregation properties that could not be rescued by PLP. Using these results, we developed a diagnostic pipeline for new patients suspected of having GACR. Adding OAT enzymatic analyses and in vitro pyridoxine responsiveness to diagnostic practices will not only increase knowledge on the consequences of pathogenic variants in OAT, but will also enable expectation management for therapeutic modalities, thus eventually improving clinical care.

A cohort study of 19 patients with gyrate atrophy of the choroid and retina (GACR).

PURPOSE: Gyrate atrophy of the choroid and retina (GACR) is an autosomal recessive inherited metabolic disorder (IMD) characterised by progressive retinal degeneration, leading to severe visual impairment. The rapid developments in ophthalmic genetic therapies warrant knowledge on clinical phenotype of eligible diseases such as GACR to define future therapeutic parameters in clinical trials. METHODS: Retrospective chart analysis was performed in nineteen patients. Data were analysed using IBM SPSS Statistics version 28.0.1.1. RESULTS: Nineteen patients were included with a mean age of 32.6 years (range 8-58). Mean age at onset of ophthalmic symptoms was 7.9 years (range 3-16). Median logMAR of visual acuity at inclusion was 0.26 (range -0.18-3.00). Mean age at cataract surgery was 28.8 years (n = 11 patients). Mean spherical equivalent of the refractive error was -8.96 (range -20.87 to -2.25). Cystoid maculopathy was present in 68% of patients, with a loss of integrity of the foveal ellipsoid zone (EZ) in 24/38 eyes. Of the 14 patients treated with dietary protein restriction, the four patients who started the diet before age 10 showed most benefit. CONCLUSION: This study demonstrates the severe ophthalmic disease course associated with GACR, as well as possible benefit of early dietary treatment. In addition to visual loss, patients experience severe myopia, early-onset cataract, and CME. There is a loss of foveal EZ integrity at a young age, emphasising the need for early diagnosis enabling current and future therapeutic interventions.

Vision on gyrate atrophy: why treat the eye?

In the April issue of this Journal, Boffa and coworkers put forward a new therapeutic approach for Gyrate Atrophy of the Choroid and Retina (GACR; OMIM 258870) (Boffa et al, 2023). The authors propose to apply gene therapy to the liver for GACR, a metabolic disease primarily affecting eyesight due to retinal degeneration. Their vision is enthusiastically supported by a News and Views comment in the same issue (Seker Yilmaz and Gissen, 2023). However, based on disease pathology, patient's needs, ethical considerations, therapeutic developmental time lines, and current state of the art of gene therapy for liver and eye, we have a different view on this issue: We argue below that local treatment of the eye is the preferred option for GACR.

A review of treatment modalities in gyrate atrophy of the choroid and retina (GACR).

Gyrate atrophy of the choroid and retina (GACR) is a rare inborn error of amino acid metabolism caused by bi-allelic variations in OAT. GACR is characterised by vision decline in early life eventually leading to complete blindness, and high plasma ornithine levels. There is no curative treatment for GACR, although several therapeutic modalities aim to slow progression of the disease by targeting different steps within the ornithine pathway. No international treatment protocol is available. We systematically collected all international literature on therapeutic interventions in GACR to provide an overview of published treatment effects. METHODS: Following the PRISMA guidelines, we conducted a systematic review of the English literature until December 22nd 2020. PubMed and Embase databases were searched for studies related to therapeutic interventions in patients with GACR. RESULTS: A total of 33 studies (n = 107 patients) met the inclusion criteria. Most studies were designed as case reports (n = 27) or case series (n = 4). No randomised controlled trials or large cohort studies were found. Treatments applied were protein-restricted diets, pyridoxine supplementation, creatine or creatine precursor supplementation, l-lysine supplementation, and proline supplementation. Protein-restricted diets lowered ornithine levels ranging from 16.0-91.2%. Pyridoxine responsiveness was reported in 30% of included mutations. Lysine supplementation decreased ornithine levels with 21-34%. Quality assessment showed low to moderate quality of the articles. CONCLUSIONS: Based primarily on case reports ornithine levels can be reduced by using a protein restricted diet, pyridoxine supplementation (variation-dependent) and/or lysine supplementation. The lack of pre-defined clinical outcome measures and structural follow-up in all included studies impeded conclusions on clinical effectiveness. Future research should be aimed at 1) Unravelling the OAT biochemical pathway to identify other possible pathologic metabolites besides ornithine, 2) Pre-defining GACR specific clinical outcome measures, and 3) Establishing an international historical cohort.