Congenital aniridia (Online Mendelian Inheritance in Man identifier, 106210) is a rare, severely visually impairing disease caused principally by heterozygous mutation in the paired box 6 (PAX6) gene that orchestrates normal ocular development. 1 The disease results in underdevelopment or abnormal development of eye structures including the cornea, leading to a bilateral and progressive limbal stem cell insufficiency and conjunctivalization of the cornea called aniridia-associated keratopathy (AAK). However, clinical manifestation of AAK, rate of progression, and prognosis can vary widely across individuals, precluding the development of general guidelines for treatment. Congenital aniridia can result from any of more than 400 unique mutations in the PAX6 gene that may lead to a spectrum of clinical phenotypes. 2 Aniridia-associated keratopathy phenotype can vary from a fully transparent cornea to a thick, opaque, vascularized pannus at any stage of life. As clinical genetic analysis becomes more sophisticated and widespread, the clinical consequence of various PAX6 mutations requires more detailed attention. However, to date, genotype–phenotype studies in aniridia describe the entire eye, 3, 4 providing only general assessment of corneal opacity. Accordingly, we performed detailed clinical characterization of AAK phenotype across a range of ages and in parallel documented PAX6 mutational status to determine how genotype influences the clinical phenotype of AAK. Adult and pediatric patients with clinically diagnosed congenital aniridia included in a patient registry maintained at the Saarland University Medical Center, Department of Ophthalmology, Homburg/Saar, Germany, were identified. Patients provided blood samples for analysis at centers in Germany specialized in clinical genetics analysis and counselling. Genetic datasets were harmonized to match with the entries of the Leiden Open Variation Database (https://www. lovd. nl/) referenced to GenBank sequence number NM_000280. 4 from the National Center for Biotechnology Information. Mutations were checked against ClinVAR, Leiden Open Variation Database, and EXAC databases and a Google search. If the mutation was not found, it was regarded as novel. Clinical examinations were conducted at the Department of Ophthalmology, Saarland University Medical Center, and consisted of slit-lamp biomicroscopy (Haag-Streit, Koeniz, Switzerland) with digital photography to perform detailed grading of AAK, Cochet-Bonnet esthesiometry (Luneau Technology, Pont-de-l’Arche, France), anterior segment swept-source OCT (Casia2; Tomey GmbH, Nürnberg, Germany) for corneal thickness measurement, and laser scanning in vivo confocal microscopy (Heidelberg Retina Tomograph 3 with Rostock Corneal Module; Heidelberg Engineering, Heidelberg, Germany) to determine central corneal epithelial phenotype and subbasal nerve density. A detailed clinical phenotypic assessment of AAK was performed for all eyes to assign an AAK grade from 0 to 4, as shown in Figure S1 (available at www. aaojournal. org). Statistical regression analysis was performed using IBM SPSS Statistics version 25 (IBM Corporation, Armonk, NY).

The collection of clinical and genetic data for this study was approved by the ethics committee of the Medical Association of Saarland (protocol no., 144/15). Written informed consent to participate was obtained from all aniridia patients (or from one or both parents of children younger than 18 years with aniridia) following the tenets of the Declaration of Helsinki. Forty-six patients in the cohort were examined bilaterally (92 eyes). The mean±standard deviation …