Challenges Using Diagnostic Next-Generation Sequencing in the Clinical Environment for Inherited Retinal Disorders

Abstract

Visual impairment, and in particular the inherited retinopathies, is a significant problem worldwide. Many disorders are progressive so their early and accurate detection is crucial to the development and application of appropriate disease management and treatment strategies, some of which are currently being tested in clinical trials. Over the past few decades, the identification of genetic causes that mediate many inherited diseases has largely been based on traditional 'Sanger sequencing' and microchip approaches that are expensive and time consuming. However, with the advent of next-generation sequencing it is now possible to apply high-throughput technologies to the clinical arena and sequence the entire exome or genome of an affected individual. Despite the potential for a paradigm shift in the clinical diagnosis of retinal disease, it may prove difficult to interpret and confirm the pathogenicity of any variants discovered by next-generation sequencing pipelines. In this review, I examine the application of next-generation sequencing to inherited retinal disorders and discuss current limitations and future perspectives.

Coupling of Human Rhodopsin to a Yeast Signaling Pathway Enables Characterization of Mutations Associated with Retinal Disease

“…Determining mutant function rapidly and accurately has become increasingly important with the rise of whole genome sequencing, and the ever-expanding rise in gene mutations with an unknown impact on human health. Rhodopsin mutations linked to inherited retinal disease have been used as examples of how many gene mutations discovered in patients are rarely characterized, and that the molecular basis for pathology is poorly understood (Davies 2014;Chiang and Gorin 2016). Animal models and traditional in vitro assays provide detailed information, but they have not kept pace with the hundreds (.350) of rhodopsin mutations identified to date.…”

Navigating the current landscape of clinical genetic testing for inherited retinal dystrophies

“…23 Several researchers have investigated the diagnostic yield of MPS technologies in patients with a variety of inherited retinal dystrophies. Using exome sequencing, Xu et al 28 found known or likely deleterious variants in 79 of 157 (50%) of families with RP, representing autosomal-dominant, autosomal-recessive, and X-linked patterns of inheritance.…”

“…Similarly, a detection rate of ~50-70% with MPS technologies was noted in several previous studies. 1,17,23 Most recently, Corton et al, 29 using exome sequencing, found causative variants in 10 of 12 families (83%), with a variety of clinical phenotypes, including RP, Usher syndrome, choroideremia, and Stargardt disease. Using NGS gene panels containing 163 genes previously known to be associated with syndromic and nonsyndromic retinopathies, Wang et al 30 found a causative mutation in 37% of 123 patients with RP.…”

“…Loss-of-function variants associated with recessive disease and dominant-negative variants, gain-of-function variants, or variants leading to haploinsufficiency have been associated with autosomal-dominant patterns of inheritance in BEST1, NR2E3 (OMIM 604485), NRL (OMIM 162080), RP1 (OMIM 603937) and RHO genes. 23,37,38 Truncating variants within the EYS gene (OMIM 612424) were initially associated with arRP in 2008. 39,40 Novel truncating variants were identified in a patient with a clinical diagnosis of autosomal-recessive CRD.…”

“…As more sequencing data become public through the efforts of data sharing initiatives, knowledge of these variants will continue to increase over time. 23,48 Laboratories performing clinical MPS testing should ensure that proper evidence is in place to classify the variant as pathogenic, such as cosegregation data, the lack of a variant in healthy family members in published reports, the absence or small representation of the variant in large control data sets (e.g., dbSNP, Exome Variant Server, and ExAc Browser), and in vitro data that show aberrant gene function. 42,[49][50][51][52][53][54][55] Many variants that have previously been reported as pathogenic in the medical literature (and thus are represented in databases used for molecular DNA analysis) are now known to have allele frequencies that make them very common occurrences in the general population, or they have been seen in the homozygous state in healthy controls.…”