Oligonucleotides targeting TCF4 triplet repeat expansion inhibit RNA foci and mis-splicing in Fuchs\u27 dystrophy

Fuchs\u27 endothelial corneal dystrophy (FECD) is the most common repeat expansion disorder. FECD impacts 4% of U.S. population and is the leading indication for corneal transplantation. Most cases are caused by an expanded intronic CUG tract in the TCF4 gene that forms nuclear foci, sequesters splicing factors and impairs splicing. We investigated the sense and antisense RNA landscape at the FECD gene and find that the sense-expanded repeat transcript is the predominant species in patient corneas. In patient tissue, sense foci number were negatively correlated with age and showed no correlation with sex. Each endothelial cell has approximately 2 sense foci and each foci is single RNA molecule. We designed antisense oligonucleotides (ASOs) to target the mutant-repetitive RNA and demonstrated potent inhibition of foci in patient-derived cells. Ex vivo treatment of FECD human corneas effectively inhibits foci and reverses pathological changes in splicing. FECD has the potential to be a model for treating many trinucleotide repeat diseases and targeting the TCF4 expansion with ASOs represents a promising therapeutic strategy to prevent and treat FECD

Identification problems with sterile fungi, illustrated by a keratitis due to a non-sporulating Chaetomium-like species

A 39-year-old farm worker was injured in her right eye by a piece of wire, which resulted in a corneal ulcer unresponsive to antibiotic treatment. The clinical appearance was that of a corneal infiltrate with feathery borders resembling fungal keratitis. Corneal scrapings were collected and the patient was started on natamycin 5% eye drops, fluconazole 0.3% eye drops, and oral fluconazole. A non-sporulating fungus was isolated from the samples. Based upon macroscopic and microscopic morphologic features, it was provisionally identified as a Papulaspora species due to the fact that members of this genus generally do not form diagnostically useful conidia. However, it was found through the use of ITS sequencing that the isolate clustered within the ascomycete genus Chaetomium. The sequence did not fully match with any sequences of available ex-type strains of Chaetomium, Thielavia and Papulaspora and hence might belong to an undescribed specie. However, without diagnostic morphological features the taxon cannot be introduced as a novel member of the genus Chaetomium. Antifungal susceptibility testing was performed according to published standards. The corneal ulcer was successfully treated with six weeks of antifungal therapy

Trinucleotide repeat expansion length as a predictor of the clinical progression of Fuchs' Endothelial Corneal Dystrophy

10.1371/journal.pone.0210996PLoS ONE141e021099

Trinucleotide repeat expansion length as a predictor of the clinical progression of Fuchs' Endothelial Corneal Dystrophy.

PURPOSE:To determine if CTG18.1 TNR expansion length prognosticates the clinical progression of Fuchs' Endothelial Corneal Dystrophy (FECD). METHODS:This was a prospective cohort study. A total of 51 patients with newly diagnosed FECD were recruited and followed-up over a period of 12 years, from November 2004 to April 2016. Baseline clinical measurements included central corneal thickness (CCT), endothelial cell density (ECD) and CTG18.1 TNR expansion length from peripheral leukocytes, with yearly repeat measurements of CCT and ECD. A patient was defined to have experienced significant clinical progression and to have developed Threshold Disease if any of these criteria were fulfilled in either eye: a) CCT increased to >700μm, b) ECD decreased to <700 cells/mm2, or c) underwent keratoplasty for treatment of FECD. RESULTS:Patients were categorized as having at least one allele whose maximum allele length was equal to or greater than 40 repeats (L≥40, n = 22, 43.1%), or having both alleles shorter than 40 repeats (L<40). Threshold Disease rates at the 5-year time point were 87.5% for the L≥40 group and 47.8% for the L<40 group (p = 0.012). This difference narrowed and was no longer statistically significant at the 8-years (92.9% vs 78.9%, p = 0.278) and 10-years (92.9% vs 84.2%, p = 0.426) time points. CONCLUSIONS:L≥40 patients are at greater risk of FECD progression and development of Threshold Disease within the first 5 years following diagnosis

Loss of Corneal Nerves and Corneal Haze in Patients with Fuchs’ Endothelial Corneal Dystrophy with the Transcription Factor 4 Gene Trinucleotide Repeat Expansion

Objective: Seventy percent of Fuchs’ endothelial corneal dystrophy (FECD) cases are caused by an intronic trinucleotide repeat expansion in the transcription factor 4 gene (TCF4). The objective of this study was to characterize the corneal subbasal nerve plexus and corneal haze in patients with FECD with (RE+) and without the trinucleotide repeat expansion (RE−) and to assess the correlation of these parameters with disease severity. Design: Cross-sectional, single-center study. Participants: Fifty-two eyes of 29 subjects with a modified Krachmer grade of FECD severity from 1 to 6 were included in the study. Fifteen of the 29 subjects carried an expanded TCF4 allele length of ≥ 40 cytosine-thymine-guanine repeats (RE+). Main Outcomes Measures: In vivo confocal microscopy assessments of corneal nerve fiber length (CNFL), corneal nerve branch density, corneal nerve fiber density (CNFD), and anterior corneal stromal backscatter (haze); Scheimpflug tomography densitometry measurements of haze in anterior, central, and posterior corneal layers. Results: Using confocal microscopy, we detected a negative correlation between FECD severity and both CNFL and CNFD in the eyes of RE+ subjects (Spearman ρ = −0.45, P = 0.029 and ρ = −0.62, P = 0.0015, respectively) but not in the eyes of RE− subjects. Additionally, CNFD negatively correlated with the repeat length of the expanded allele in the RE+ subjects (Spearman ρ = −0.42, P = 0.038). We found a positive correlation between anterior stromal backscatter and severity in both the RE+ and RE− groups (ρ = 0.60, P = 0.0023 and ρ = 0.44, P = 0.024, respectively). The anterior, central, and posterior Scheimpflug densitometry measurements also positively correlated with severity in both the RE+ and RE− groups (P = 5.5 × 10−5, 2.5 × 10−4, and 2.9 × 10−4, respectively, after adjusting for the expansion status in a pooled analysis. However, for patients with severe FECD (Krachmer grades 5 and 6), the posterior densitometry measurements were higher in the RE+ group than in the RE− group (P < 0.05). Conclusions: Loss of corneal nerves in FECD supports the classification of the TCF4 trinucleotide repeat expansion disorder as a neurodegenerative disease. Haze in the anterior, central, and posterior cornea correlate with severity, irrespective of the genotype. Quantitative assessments of corneal nerves and corneal haze may be useful to gauge and monitor FECD disease severity in RE+ patients