Mutation Analysis of Japanese Patients with Leber Congenital Amaurosis by Next Generation Sequencing

Abstract

Purpose : Leber congenital amaurosis (LCA), a genetically and clinically heterogeneous disease, is a severe form of inherited retinal dystrophies (IRD). We performed the mutation analysis in 34 Japanese families with LCA to evaluate the mutation spectrum and frequency of known LCA-associated genes in the Japanese population.

Methods : Patients who fulfilled the following criteria were included in the study: 1. severe visual impairment found within the first year after birth, 2. presence of nystagmus and poor visual response, 3. severely reduced or non-detectable rod and cone electroretinography, 4.maximum best-corrected decimal visual acuity ≤ 0.5, and 5. absence of systemic abnormality other than neurodevelopmental delay. Clinical analyses were based on ophthalmic examinations, fundus photography, and electroretinography. Seventy-four genes responsible for IRD were examined by targeted-next generation sequencing (NGS) using a custom designed Agilent HaloPlex target enrichment kit with Illumina Miseq sequencer to identify potential pathogenic mutations. The identified potential pathogenic mutations were confirmed using Sanger sequencing. Segregation analyses were performed on DNA from available family members to investigate the co-segregation of potentially pathogenic mutations. Quantitative real-time polymerase chain reaction (qPCR) was used to screen copy number variations in some patients in whom the targeted NGS approach revealed a single heterozygous mutations in autosomal recessive LCA-associated genes.

Results : Twenty-nine potential pathogenic mutations were identified in 18 out of 34 families in 11 genes (nine LCA-associated genes and two IRD-associated genes). The frequently mutated genes were CRB1, NMNAT1, and RPGRIP1. qPCR detected a LCA family with a large heterozygous deletion mutation including exons 1–3 of LRAT. The mutation spectrum and frequency in the Japanese population is distinctly different from that in the European population.

Conclusions : We reported the results of the first NGS-based molecular diagnosis of the largest Japanese LCA cohort so far. The observed detection rate of about 53% indicated that targeted NGS is an effective tool for the diagnosis of LCA.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.