Copy number variation - missing inheritance in retinal disorder

Abstract

Purpose : Inherited retinal degenerations (IRDs) are an important cause of blindness affecting over two million people worldwide. Even though IRDs are mostly monogenic, they are genetically heterogeneous with mutations in over 200 genes leading to disease. Despite substantial progress in sequencing and new disease gene discovery, current strategies can identify the genetic causes for only about 60% of IRD cases. The high number of unsolved cases can be attributed to the yet-unidentified genes, large duplications/deletions also called copy number variations (CNVs), and deep intronic mutations, which are not detected by targeted NGS approaches. The purpose of this study was to screen genetically unsolved patients for CNVs.

Methods : The patients included in the study were recruited and clinically examined at the Massachusetts Eye and Ear Infirmary and/or The Children’s Hospital of Philadelphia. Patients underwent a full ophthalmic examination and their DNA samples were studied by Genetic Eye Disease (GEDi) diagnostic test or whole exome sequencing (WES), SNP genotyping array, comparative genomic hybridization array, quantitative real-time PCR (qRT-PCR), PCR and Sanger sequencing. The study protocol adhered to the tenets of the Declaration of Helsinki and was approved by the Institutional Review Boards. All subjects signed informed consent.

Results : Twenty six families previously unsolved with WES and two families with apparently homozygous mutations in known IRD genes were studied for CNVs. We detected likely pathogenic deletions in five families. These included: a heterozygous ≈1.1 Mb deletion on chromosome 2, affecting 20 genes including SNRNP200 and CNNM4 in two families (in trans with c.3133C>A, p.P1045T in SNRNP200 and c.480G>A, p.W160* in CNNM4 respectively), a heterozygous deletion of ≈34kb affecting PRPF31, a heterozygous deletion of exons 15-18 in EYS (in trans with c.9099_9099del, p.G3033fs) and a hemizygous deletion of 52.6 kb affecting the Opsin genes locus control region, the entire red opsin gene (OPN1LW) and the first exon of the green opsin gene (OPN1MW). All the deletions were confirmed and fine mapped by qRT-PCR. In two families, the breakpoints were determined by PCR amplification across the breakpoints and direct sequencing.

Conclusions : The genetic cause of five out of 28 IRD families could be attributed to CNVs (18%), which demonstrates that CNVs have a considerable contribution to the etiology of IRDs.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.