Vincent Soler

Designing dendrimers for ocular drug delivery

New series of phosphorus-containing dendrimers, having one quaternary ammonium salt as core and carboxylic acid terminal groups have been synthesized from generation 0 (3 carboxylic acid terminal groups) to generation 2 (12 carboxylic acid terminal groups). These dendrimers react with the neutral form of carteolol (an ocular anti-hypertensive drug used to treat glaucoma) to afford ion pair (saline) species. The solubility in water of these charged dendrimers depends on the generation considered: generation 0 (3 carteolol) is well soluble, whereas generation 1 (6 carteolol) and generation 2 (12 carteolol) are poorly soluble. These dendrimers have been tested in vivo, as vehicle for ocular drug delivery of carteolol to rabbits.

Mutations in SCO2 Are Associated with Autosomal-Dominant High-Grade Myopia

Myopia, or near-sightedness, is an ocular refractive error of unfocused image quality in front of the retinal plane. Individuals with high-grade myopia (dioptric power greater than -6.00) are predisposed to ocular morbidities such as glaucoma, retinal detachment, and myopic maculopathy. Nonsyndromic, high-grade myopia is highly heritable, and to date multiple gene loci have been reported. We performed exome sequencing in 4 individuals from an 11-member family of European descent from the United States. Affected individuals had a mean dioptric spherical equivalent of -22.00 sphere. A premature stop codon mutation c.157C>T (p.Gln53*) cosegregating with disease was discovered within SCO2 that maps to chromosome 22q13.33. Subsequent analyses identified three additional mutations in three highly myopic unrelated individuals (c.341G>A, c.418G>A, and c.776C>T). To determine differential gene expression in a developmental mouse model, we induced myopia by applying a -15.00D lens over one eye. Messenger RNA levels of SCO2 were significantly downregulated in myopic mouse retinae. Immunohistochemistry in mouse eyes confirmed SCO2 protein localization in retina, retinal pigment epithelium, and sclera. SCO2 encodes for a copper homeostasis protein influential in mitochondrial cytochrome c oxidase activity. Copper deficiencies have been linked with photoreceptor loss and myopia with increased scleral wall elasticity. Retinal thinning has been reported with an SC02 variant. Human mutation identification with support from an induced myopic animal provides biological insights of myopic development.

Iontophoresis Transcorneal Delivery Technique for Transepithelial Corneal Collagen Crosslinking With Riboflavin in a Rabbit Model.

PURPOSE We compared an iontophoresis riboflavin delivery technique for transepithelial corneal collagen crosslinking (I-CXL) with a conventional CXL (C-CXL). METHODS We designed three experimental sets using 152 New Zealand rabbits to study riboflavin application by iontophoresis using charged riboflavin solution (Ricrolin+) with a 1-mA current for 5 minutes. The first set was to compare riboflavin concentration measured by HPLC in corneas after iontophoresis or conventional riboflavin application. The second set was to analyze autofluorescence and stromal collagen modification immediately and 14 days after I-CXL or C-CXL, by using nonlinear two-photon microscopy (TP) and second harmonic generation (SHG). In the third set, physical modifications after I-CXL and C-CXL were evaluated by stress-strain measurements and by studying corneal resistance against collagenase digestion. RESULTS Based on HPLC analysis, we found that iontophoresis allowed riboflavin diffusion with 2-fold less riboflavin concentration than conventional application (936.2 ± 312.5 and 1708 ± 908.3 ng/mL, respectively, P < 0.05). Corneal TP and SHG imaging revealed that I-CXL and C-CXL resulted in a comparable increased anterior and median stromal autofluorescence and collagen packing. The stress at 10% strain showed a similar stiffness of corneas treated by I-CXL or C-CXL (631.9 ± 241.5 and 680.3 ± 216.4 kPa, respectively, P = 0.908). Moreover, we observed an increased resistance against corneal collagenase digestion after I-CXL and C-CXL (61.90% ± 5.28% and 72.21% ± 4.32% of remaining surface, respectively, P = 0.154). CONCLUSIONS This experimental study suggests that I-CXL is a promising alternative methodology for riboflavin delivery in crosslinking treatments, preserving the epithelium.

Whole exome sequencing identifies a mutation for a novel form of corneal intraepithelial dyskeratosis

Background Corneal intraepithelial dyskeratosis is an extremely rare condition. The classical form, affecting Native American Haliwa-Saponi tribe members, is called hereditary benign intraepithelial dyskeratosis (HBID). Herein, we present a new form of corneal intraepithelial dyskeratosis for which we identified the causative gene by using deep sequencing technology. Methods and results A seven member Caucasian French family with two corneal intraepithelial dyskeratosis affected individuals (6-year-old proband and his mother) was ascertained. The proband presented with bilateral complete corneal opacification and dyskeratosis. Palmoplantar hyperkeratosis and laryngeal dyskeratosis were associated with the phenotype. Histopathology studies of cornea and vocal cord biopsies showed dyskeratotic keratinisation. Quantitative PCR ruled out 4q35 duplication, classically described in HBID cases. Next generation sequencing with mean coverage of 50× using the Illumina Hi Seq and whole exome capture processing was performed. Sequence reads were aligned, and screened for single nucleotide variants and insertion/deletion calls. In-house pipeline filtering analyses and comparisons with available databases were performed. A novel missense mutation M77T was discovered for the gene NLRP1 which maps to chromosome 17p13.2. This was a de novo mutation in the probands mother, following segregation in the family, and not found in 738 control DNA samples. NLRP1 expression was determined in adult corneal epithelium. The amino acid change was found to destabilise significantly the protein structure. Conclusions We describe a new corneal intraepithelial dyskeratosis and how we identified its causative gene. The NLRP1 gene product is implicated in inflammation, autoimmune disorders, and caspase mediated apoptosis. NLRP1 polymorphisms are associated with various diseases.

Risk Factors for Progressive Myopia in the Atropine Therapy for Myopia Study

PURPOSE To investigate variables associated with myopic progression despite treatment in the Atropine in the Treatment of Myopia Study. DESIGN Retrospective cohort study. METHODS Two hundred of 400 children were randomized to receive atropine 1% in 1 eye only in this institutional study. Children were followed up with cycloplegic autorefraction every 4 months over 2 years. Children whose myopia progressed by more than 0.5 diopter (D) in the atropine-treated eye at 1 year were classified as being progressors. RESULTS Among the 182 children still in the study at 1 year, 22 (12.1%) were classified as progressors. Univariate analysis suggested these children tended to be younger (8.5 ± 1.4 years vs 9.3 ± 1.5 years; P = .023), to have higher myopic spherical equivalent (SE) at baseline (-3.6 ± 1.3 D vs -2.8 ± 1.4 D; P = .015), and to have 2 myopic parents (77.3% vs 48.1%; P = .012). In nonprogressors, the myopia progression at 1 year was less in the atropine-treated eyes compared with the untreated fellow eye (+0.16 ± 0.37 D vs -0.73 ± 0.48 D; P < .001), but in progressors, progression was more similar between eyes (-0.92 ± 0.31 D vs -1.06 ± 0.44 D; P = .363). Regression analysis showed that the risk of being a progressor was 40% lower with each year of increased age, 43% lower for every 1.0 D less in myopia at baseline, and 59% lower for every 1.0 D less in myopic change in the untreated eyes over the first year. CONCLUSIONS Doctors and parents need to be aware that there is a small group of children (younger, with higher myopia, and greater tendency of myopic progression) who may still progress while receiving atropine treatment.

A randomized comparison of pupil-centered versus vertex-centered ablation in LASIK correction of hyperopia.

PURPOSE To compare visual and optical outcomes of pupil-centered vs vertex-centered ablation in patients undergoing laser-assisted in situ keratomileusis (LASIK) for hyperopia. DESIGN Randomized, double-masked, prospective, single-center trial. METHODS SETTING Institutional practice. STUDY POPULATION Sixty eyes of 30 patients with low and moderate hyperopia. Intervention procedure: Eyes underwent LASIK (Allegretto excimer laser). In 30 eyes, the ablation was centered on the pupil, while in the 30 other eyes the ablation was centered on the corneal reflex. MAIN OUTCOME MEASURES Primary outcome measure was the safety index. Main secondary outcome measures were efficacy index, manifest refraction, uncorrected visual acuity, best spectacle-corrected visual acuity (BCVA), and ocular high-order aberrations for a 6-mm pupil size. RESULTS At 3 months postoperatively, the safety index was 0.99 ± 0.04 in the pupil-centered group and 0.99 ± 0.08 in the vertex-centered group (P = .97). The efficacy index was also similar for both groups: 0.96 ± 0.05 in pupil-centered eyes and 0.93 ± 0.09 in vertex-centered eyes (P = .31). Optical aberrations were similar for pupil-centered and vertex-centered eyes. Considering only eyes showing large pupil decentration, we found a tendency for better visual results in favor of pupil-centered eyes in terms of safety index and a slight but significant increase of coma in vertex-centered eyes. CONCLUSION LASIK is an effective procedure for treatment of hyperopia. Pupil-centered and vertex-centered treatments provide similar visual and optical outcomes. However, in eyes showing large temporal pupil decentration, pupil-centered ablation seemed to produce a lower amount of coma and, as a consequence, a reduced loss of BCVA compared with vertex-centered patients.

Reduced penetrance in a large Caucasian pedigree with Stickler syndrome.

ABSTRACT Background: In a four-generation Caucasian family variably diagnosed with autosomal dominant (AD) Stickler or Wagner disease, commercial gene screening failed to identify a mutation in COL2A1 or VCAN. We utilized linkage mapping and exome sequencing to identify the causal variant. Materials and methods: Genomic DNA samples collected from 40 family members were analyzed. A whole-genome linkage scan was performed using Illumina HumanLinkage-24 BeadChip followed by two-point and multipoint linkage analyses using FASTLINK and MERLIN. Exome sequencing was performed on two affected individuals, followed by co-segregation analysis. Results: Parametric multipoint linkage analysis using an AD inheritance model demonstrated HLOD scores > 2.00 at chromosomes 1p36.13-1p36.11 and 12q12-12q14.1. SIMWALK multipoint analysis replicated the peak in chromosome 12q (peak LOD = 1.975). FASTLINK two-point analysis highlighted several clustered chromosome 12q SNPs with HLOD > 1.0. Exome sequencing revealed a novel nonsense mutation (c.115C>T, p.Gln39*) in exon 2 of COL2A1 that is expected to result in nonsense-mediated decay of the RNA transcript. This mutation co-segregated with all clinically affected individuals and seven individuals who were clinically unaffected. Conclusions: The utility of combining traditional linkage mapping and exome sequencing is highlighted to identify gene mutations in large families displaying a Mendelian inheritance of disease. Historically, nonsense mutations in exon 2 of COL2A1 have been reported to cause a fully penetrant ocular-only Stickler phenotype with few or no systemic manifestations. We report a novel nonsense mutation in exon 2 of COL2A1 that displays incomplete penetrance and/or variable age of onset with extraocular manifestations.

First Experience With the ICD 16.5 Mini-Scleral Lens for Optic and Therapeutic Purposes.

Objectives: To evaluate the success rate, efficacy, and safety of the ICD 16.5 mini-scleral gas permeable (GP) contact lens. Methods: This prospective study included referred consecutive patients with irregular corneas and severe ocular surface disease (OSD) in treatment failure. All patients were fitted with the ICD 16.5 mini-scleral GP lens. Even though we had some limited experience with scleral lenses, it was our first experience with the ICD 16.5 mini-scleral GP lens. Efficacy was assessed by comparing best-corrected visual acuity (BCVA) with the mini-scleral lens to baseline BCVA. A subjective visual functioning questionnaire (comfort score, visual quality score, handling rating, and wearing time) was administered in a face-to-face structured interview. Results: Thirty-nine eyes of 23 patients with a mean age of 43±16 years were included. Fitting indications were keratoconus (46%), post-penetrating keratoplasty (21%), other irregular astigmatism (15%), and severe OSD (18%). Twenty-five eyes (64%) were successfully fitted with an 18-month follow-up. The mini-scleral GP lens BCVA was 0.16 logarithm of the minimum angle of resolution (logMAR; 20/25) versus a baseline BCVA of 0.44 logMAR (20/63; P<0.001). Comfort and visual quality scores were 8.5/10 and 7.5/10, respectively. No complications were detected in 96% of the eyes (95% confidence interval, 76.1%–99.4%). One eye experienced corneal graft swelling. Conclusions: The present findings suggest that the ICD 16.5 mini-scleral GP lens is an effective and safe alternative for managing challenging corneas in a therapeutic impasse.

Five-year follow-up of bilateral choroidal neovascularization secondary to optic nerve head drusen treated with ranibizumab in a nine-year-old girl.

We report the case of a nine-year-old girl who was referred to L’Union Private Hospital (Saint-Jean, Midi-Pyrénées, France) in May 2009 for a recent visual impairment in the left eye secondary to sub-retinal neovascularization associated with optic nerve head drusen (ONHD). She had no relevant ocular or general history. Visual acuity was limited to 20/200 in the left eye, and was 20/20 in the right eye. Anterior segment and intraocular pressure were normal in both eyes. Fundus examination revealed a bilateral lumpy aspect of the ONH (Fig. 1A/B), which were hyper-autofluorescent leading to the diagnosis of bilateral ONHD (Fig. 1C/D). In the left eye, there was an inter-papillo-macular and elevated grey lesion (Fig. 1B), which showed early hyperfluorescence, and an intense leakage at fluorescein angiography (Fig. 1F). OCT scans showed retinal thickening, sub-retinal hyperreflectivity in the interpapillo-macular region and sub-retinal fluid in the subfoveal area (Fig. 2A). These findings lead to the diagnosis of choroidal neovascular membrane secondary to ONHD in the left eye. A single ranibizumab (0.5 mg/0.05 mL) intravitreal injection was administered. One month later, visual acuity was limited to 20/200 and OCT scan showed the decrease of the retinal thickening and the persistence of sub-retinal hyperreflectivity (Fig. 2B). At the two-month follow-up examination, retinal thickening increased anew. The patient underwent a second ranibizumab intravitreal injection in the left eye. The five-month examination revealed visual acuity improvement at 20/40; OCT showed anatomical improvement with the persistence of sub-retinal fibrosis. In May 2010, the patient presented with a recent visual impairment in the right eye which visual acuity was 20/25. Fundus examination, fluorescein angiography (Fig. 3) and OCT scans showed similar lesions in the right eye as the ones observed in the left eye one year before. The patient underwent a third ranibizumab intravitreal injection (the first in the right eye). Follow-up examinations showed functional a b

Genetics of keratoconus

Keratoconus (KC) is a vision‐threatening condition characterized by thinning and deformation of the cornea. It is one of the most common indications for corneal grafting in industrialized countries. The disease prevalence is approximately 1 in 2000. Familial aggregation, together with increased familial risk, suggests important genetic influences on its pathogenesis. To date, it has been shown that several genomic regions are linked to KC rare familial forms but no genes have been identified as responsible for common KC. Aside from genetic determinants, environmental stresses such as eye rubbing or atopy have been suggested as possible causes or aggravating factors in KC. The interaction of genetics and environmental factors in diseases occurrence contributes to the development of complex and multifactorial trait, such as KC. And, despite the many attempts to reveal KC pathophysiology, the mechanisms leading to its corneal characteristics and vision impairment are still poorly understood. In this presentation, we will discuss all works that have been achieved on keratoconus pathophysiology and we will consider future directions on this topic.