Prevalence of FOXC1 Variants in Individuals With a Suspected Diagnosis of Primary Congenital Glaucoma

Abstract

Importance: Both primary and secondary forms of childhood glaucoma have many distinct causative mechanisms, and in many cases a cause is not immediately clear. The broad phenotypic spectrum of secondary glaucoma, particularly in individuals with variants in FOXC1 or PITX2 genes associated with Axenfeld-Rieger syndrome, makes it more difficult to diagnose patients with milder phenotypes. These cases are occasionally classified and managed as primary congenital glaucoma.

Objective: To investigate the prevalence of FOXC1 variants in participants with a suspected diagnosis of primary congenital glaucoma.

Design, setting, and participants: Australian and Italian cohorts were recruited from January 1, 2007, through March 1, 2016. Australian individuals were recruited through the Australian and New Zealand Registry of Advanced Glaucoma and Italian individuals through the Genetic and Ophthalmology Unit of l'Azienda Socio-Sanitaria Territoriale Grande Ospedale Metropolitano Niguarda in Milan, Italy. We performed exome sequencing, in combination with Sanger sequencing and multiplex ligation-dependent probe amplification, to detect variants of FOXC1 in individuals with a suspected diagnosis of primary congenital glaucoma established by their treating specialist. Data analysis was completed from June 2015 to November 2017.

Main outcome and measures: Identification of single-nucleotide and copy number variants in FOXC1, along with phenotypic characterization of the individuals who carried them.

Results: A total of 131 individuals with a suspected diagnosis of primary congenital glaucoma were included. The mean (SD) age at recruitment in the Australian cohort was 24.3 (18.1) years; 37 of 84 Australian participants (44.0%) were female, and 71 of 84 (84.5%) were of European ancestry. The mean (SD) age at recruitment was 22.5 (18.4) years in the Italian cohort; 21 of 47 Italian participants (44.7%) were female, and 45 of 47 (95.7%) were of European ancestry. We observed rare, predicted deleterious FOXC1 variants in 8 of 131 participants (6.1%), or 8 of 166 participants (4.8%) when including those explained by variants in CYP1B1. On reexamination or reinvestigation, all of these individuals had at least 1 detectable ocular and/or systemic feature associated with Axenfeld-Rieger syndrome.

Conclusions and relevance: These data highlight the genetic and phenotypic heterogeneity of childhood glaucoma and support the use of gene panels incorporating FOXC1 as a diagnostic aid, especially because clinical features of Axenfeld-Rieger syndrome can be subtle. Further replication of these results will be needed to support the future use of such panels.

Figure 1.. FOXC1 Variants Associated With Suspected Primary Congenital Glaucoma (PCG)

A, Frameshift and missense variants identified by exome sequencing. B, Exome sequencing read depth at the FOXC1 locus (chr6:1390204-2679449, hg19) for participant PCG029 compared with the mean read depth across all suspected PCG samples. C, Multiplex ligation-dependent probe amplification (MLPA) using probes within FOXC1 (chr6) and on both sex chromosomes. Signal intensity was measured in duplicate, normalized, and plotted as the mean for participant PCG029 and all other samples processed in the same batch (control participants). D, Mean coverage across the FOXC1 locus for exomes (gray) and genomes (black) from the gnomAD release 2.0.2 collection, indicating location of variants identified by exome (blue fill) or capillary (no fill) sequencing. Multiplex ligation-dependent probe amplification probe locations are indicated in red. Most gnomAD exome sequences (approximately 77%) were processed with the same or similar exome capture kits as those used in the current study (Agilent SureSelect).

Figure 2.. Segregation of FOXC1 Variants

A, Capillary sequencing trace images. B, Location of missense, nonsense, frameshift, and nonframeshift deletion variants within the FOXC1 protein; the forkhead domain is indicated by black shading.