September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Molecular inversion probe based sequence analysis of 108 genes associated with non-syndromic inherited retinal disease in 4,000 probands
Author Affiliations & Notes
  • M Imran Khan
    Department of Human Genetics, Radboud university medical centre, Nijmegen, Netherlands
  • Gilissen Christian
    Department of Human Genetics, Radboud university medical centre, Nijmegen, Netherlands
  • Ayuso Carmen
    Department of Genetics, University Hospital Fundación Jiménez Díaz , Madrid, Spain
  • Dror Sharon
    Departments of Ophthalmology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
  • Robert K Koenekoop
    Departments of Paediatric Surgery, Human Genetics and Ophthalmology, McGill University Health Center, Montreal, Quebec, Canada
  • Carlo Rivolta
    Department of Medical Genetics, University of Lausanne, Lausanne, Switzerland
  • Elfride De Baere
    Center for Medical Genetics, Ghent University Hospital, Ghent, Belgium
  • Chris F Inglehearn
    Department of Ophthalmology and Neuroscience, University of Leeds, Leeds, United Kingdom
  • Alexander Hoischen
    Department of Human Genetics, Radboud university medical centre, Nijmegen, Netherlands
  • Frans P Cremers
    Department of Human Genetics, Radboud university medical centre, Nijmegen, Netherlands
    Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, Netherlands
  • Footnotes
    Commercial Relationships   M Imran Khan, None; Gilissen Christian, None; Ayuso Carmen, None; Dror Sharon, None; Robert Koenekoop, None; Carlo Rivolta, None; Elfride De Baere, None; Chris Inglehearn, None; Alexander Hoischen, None; Frans Cremers, None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science September 2016, Vol.57, No Pagination Specified. doi:
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      M Imran Khan, Gilissen Christian, Ayuso Carmen, Dror Sharon, Robert K Koenekoop, Carlo Rivolta, Elfride De Baere, Chris F Inglehearn, Alexander Hoischen, Frans P Cremers; Molecular inversion probe based sequence analysis of 108 genes associated with non-syndromic inherited retinal disease in 4,000 probands. Invest. Ophthalmol. Vis. Sci. 201657(12):.

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      © 2017 Association for Research in Vision and Ophthalmology.

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Abstract

Purpose : Inherited retinal diseases (IRDs) are clinically and genetically very heterogeneous, as ~125 genes have been associated with non-syndromic IRDs. The purpose of this study was to develop a flexible, comprehensive and cost effective sequencing procedure for IRDs.

Methods : In total, 6,200 molecular inversion probes (MIPs) were designed to capture ~1600 protein-coding exons and flanking intronic sequences of 108 IRD-associated genes published up to October 2013. About 4,000 probands with non-syndromic IRDs were ascertained by partners of the European Retinal Disease Consortium (ERDC). The most prevalent phenotypes were retinitis pigmentosa, cone-rod dystrophy, and Leber congenital amaurosis. To analyze the performance of the MIPs, three tests were performed. The captured targets were sequenced on a NextSeq500 Illumina sequencer and the data were analyzed using an in-house pipeline to find the causal genetic variants. The over- and poor-performing probes were rebalanced and used to capture the genomic targets in 4,000 individuals.

Results : In pools of 120 samples the average coverage per probe was ~500X, where 95.6% of the probes were covered >10X. Probes targeting 272 (4.4%) regions were either poorly covered (2.3%) or not covered (2.1%). Analysis was completed for 2,500/4,000 samples. Sanger validation was performed for a representative set of variants identified in 290 probands from Nijmegen. Material costs (including MIPs synthesis, library preparations and sequencing costs) for sequencing 108 IRD genes, was € 65 per sample, which makes it very cost-effective. We identified the causal variants in 59% (172/290) of the cases, 36 of 118 unsolved cases (i.e. 12% of total) carry one likely pathogenic variant in an autosomal recessive gene.

Conclusions : Taken into consideration that the Nijmegen cohort was previously prescreened using various genotyping methods, the corrected yield would be ~71%. At 1/10th of WES cost, this efficiency is equal or superior to other published gene-panel (36 - 62%) or WES-based (49 - 66%) sequence analysis. Based on preliminary results, we estimate to identify causal variants in at least 2,000 IRD probands.

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

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