Mutation Detection in Patients with Retinal Dystrophies Using Targeted Next Generation Sequencing

RESEARCH ARTICLE Mutation Detection in Patients with Retinal Dystrophies Using Targeted Next Generation Sequencing Nicole Weisschuh1*, Anja K. Mayer1, Tim M. Strom2, Susanne Kohl1, Nicola Glöckle3, Max Schubach4, Sten Andreasson5, Antje Bernd6, David G. Birch7, Christian P. Hamel8, John R. Heckenlively9, Samuel G. Jacobson10, Christina Kamme5, Ulrich Kellner11, Erdmute Kunstmann12, Pietro Maffei13, Charlotte M. Reiff14, Klaus Rohrschneider15, Thomas Rosenberg16, Günther Rudolph17, Rita Vámos18, Balázs Varsányi18,19, Richard G. Weleber20, Bernd Wissinger1 OPEN ACCESS Citation: Weisschuh N, Mayer AK, Strom TM, Kohl S, Glöckle N, Schubach M, et al. (2016) Mutation Detection in Patients with Retinal Dystrophies Using Targeted Next Generation Sequencing. PLoS ONE 11(1): e0145951. doi:10.1371/journal.pone.0145951 Editor: Andreas R. Janecke, Innsbruck Medical University, AUSTRIA 1 Molecular Genetics Laboratory, Institute for Ophthalmic Research, Centre for Ophthalmology, University of Tuebingen, Tuebingen, Germany, 2 Institute of Human Genetics, Helmholtz Zentrum Muenchen, Neuherberg, Germany, 3 CeGaT GmbH, Tuebingen, Germany, 4 Institute of Medical Genetics and Human Genetics, Charité – Universitaetsmedizin Berlin, Berlin, Germany, 5 Department of Ophthalmology, Lund University, Lund, Sweden, 6 University Eye Hospital, Centre for Ophthalmology, University of Tuebingen, Tuebingen, Germany, 7 The Retina Foundation of the Southwest, Dallas, Texas, United States of America, 8 Genetic Sensory Diseases, CHU de Montpellier, Montpellier, France, 9 Department of Ophthalmology and Visual Sciences, Kellogg Eye Center, University of Michigan, Ann Arbor, Michigan, United States of America, 10 Scheie Eye Institute, Department of Ophthalmology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America, 11 Rare Retinal Disease Center, AugenZentrum Siegburg, MVZ ADTC Siegburg GmbH, Siegburg, Germany, 12 Institute of Human Genetics, Julius-Maximilian-University, Wuerzburg, Germany, 13 Department of Medicine, University Hospital of Padua, Padua, Italy, 14 Eye Center, Albert-Ludwigs-University of Freiburg, Freiburg, Germany, 15 Department of Ophthalmology, University of Heidelberg, Heidelberg, Germany, 16 National Eye Clinic, Department of Ophthalmology, Glostrup Hospital, Glostrup, Denmark, 17 University Eye Hospital, Ludwig Maximilians University, Munich, Germany, 18 Department of Ophthalmology, Semmelweis University, Budapest, Hungary, 19 Department of Ophthalmology, University of Pécs Medical School, Pécs, Hungary, 20 Casey Eye Institute, Oregon Retinal Degeneration Center, Oregon Health & Science University, Portland, Oregon, United States of America Received: September 9, 2015 Accepted: December 10, 2015 * Published: January 14, 2016 Copyright: © 2016 Weisschuh et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: All relevant data are within the paper. Funding: This work was supported through grants (01GM1105A to BW and 01GM1108A to BW and SK) from the German Ministry for Education and Research, http://www.bmbf.de/en/. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: NG is an employee of CeGaT GmbH. There are no patents, products in development or marketed products to declare. This Abstract Retinal dystrophies (RD) constitute a group of blinding diseases that are characterized by clinical variability and pronounced genetic heterogeneity. The different nonsyndromic and syndromic forms of RD can be attributed to mutations in more than 200 genes. Consequently, next generation sequencing (NGS) technologies are among the most promising approaches to identify mutations in RD. We screened a large cohort of patients comprising 89 independent cases and families with various subforms of RD applying different NGS platforms. While mutation screening in 50 cases was performed using a RD gene capture panel, 47 cases were analyzed using whole exome sequencing. One family was analyzed using whole genome sequencing. A detection rate of 61% was achieved including mutations in 34 known and two novel RD genes. A total of 69 distinct mutations were identified, including 39 novel mutations. Notably, genetic findings in several families were not consistent with the initial clinical diagnosis. Clinical reassessment resulted in refinement of the PLOS ONE | DOI:10.1371/journal.pone.0145951 January 14, 2016 1 / 15 Mutation Detection in RD Patients Using Targeted NGS does not alter the authors' adherence to PLOS ONE policies on sharing data and materials. clinical diagnosis in some of these families and confirmed the broad clinical spectrum associated with mutations in RD genes. Introduction Retinal dystrophies (RD) are among the disorders with the highest level of heterogeneity. This includes genetic heterogeneity, allelic heterogeneity as well as clinical heterogeneity. Molecular genetic studies in the last two decades revealed ~225 genes that are mutated in one or more of the various clinical subtypes of RD (https://sph.uth.edu/retnet/). Some of the clinical subtypes of RD can be caused by mutations in up to 60 different genes, e.g. in retinitis pigmentosa (RP). Adding to the genetic complexity there is considerable variation in clinical expression and overlap of symptoms of single disease entities, all of which may hamper making an exact clinical diagnosis. These obstacles have also practical implications for molecular diagnostics. Because it is difficult to predict the gene likely to be mutated, a gene-by-gene screening approach in RD patients is neither time- nor cost-efficient. On the other hand, establishing a molecular diagnosis is important for several reasons. It is vital for determining the recurrence risk for future children and therefore provides the basis for accurate genetic counseling. In many instances, it will also help to predict the clinical course, which is of central importance for the patients to plan and organize their professional and social lives. There is no effective cure for RD, however, ongoing clinical trials applying gene-replacement therapy approaches for several forms of RD have raised new hopes. Since these approaches require the identification of the causative mutation, the genetic diagnosis is an essential prerequisite. The identification of the genetic defect in RD patients has been accelerated by the introduction of next-generation-sequencing technologies (NGS). Within the field of NGS platforms, the targeted capture of known disease genes (“disease panels”) has been proven superior in terms of coverage compared with whole exome sequencing (WES), especially for previous generations of exome capture (...truncated)