June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Single-molecule Molecular Inversion Probes allow cost-effective targeted sequencing of all genes and loci associated with macular diseases.
Author Affiliations & Notes
  • Rebekkah J Hitti-Malin
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Radboud Universiteit Donders Institute for Brain Cognition and Behaviour, Nijmegen, Gelderland, Netherlands
  • Susanne Roosing
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Radboud Universiteit Donders Institute for Brain Cognition and Behaviour, Nijmegen, Gelderland, Netherlands
  • Claire-Marie Dhaenens
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Lille Neuroscience & Cognition, Universite de Lille Faculte de Medecine, Lille, Hauts-de-France, France
  • Anneke I Den Hollander
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Department of Ophthalmology, Radboudumc, Nijmegen, Gelderland, Netherlands
  • G.Jane Farrar
    The School of Genetics & Microbiology, The University of Dublin Trinity College, Dublin, Ireland
  • Daan Panneman
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Radboud Universiteit Donders Institute for Brain Cognition and Behaviour, Nijmegen, Gelderland, Netherlands
  • Erica G.M Boonen
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
  • Laura de Rooij
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
  • Mariana Guimaraes Ramos
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
  • Maartje van de Vorst
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
  • Bart de Koning
    Department of Clinical Genetics, Maastricht Universitair Medisch Centrum+, Maastricht, Limburg, Netherlands
  • Christian Gilissen
    Radboud Institute of Molecular Life Sciences, Radboudumc, Nijmegen, Gelderland, Netherlands
  • Alexander Hoischen
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Radboud Institute of Molecular Life Sciences, Radboudumc, Nijmegen, Gelderland, Netherlands
  • Frans P Cremers
    Department of Human Genetics, Radboudumc, Nijmegen, Gelderland, Netherlands
    Radboud Universiteit Donders Institute for Brain Cognition and Behaviour, Nijmegen, Gelderland, Netherlands
  • Footnotes
    Commercial Relationships   Rebekkah Hitti-Malin None; Susanne Roosing Novartis, Code F (Financial Support); Claire-Marie Dhaenens None; Anneke Den Hollander AbbVie, Code E (Employment); G.Jane Farrar None; Daan Panneman Novartis, Code F (Financial Support); Erica Boonen Novartis, Code F (Financial Support); Laura de Rooij None; Mariana Guimaraes Ramos None; Maartje van de Vorst None; Bart de Koning None; Christian Gilissen None; Alexander Hoischen None; Frans Cremers Novartis, Code F (Financial Support)
  • Footnotes
    Support  HRCI-HRB 2020, Oogfonds, Pro Retina Deutschland, Stichting ter Verbetering van het Lot der Blinden, Stichting voor Ooglijders, Stichting Blindenhulp
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 41. doi:
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    • Get Citation

      Rebekkah J Hitti-Malin, Susanne Roosing, Claire-Marie Dhaenens, Anneke I Den Hollander, G.Jane Farrar, Daan Panneman, Erica G.M Boonen, Laura de Rooij, Mariana Guimaraes Ramos, Maartje van de Vorst, Bart de Koning, Christian Gilissen, Alexander Hoischen, Frans P Cremers; Single-molecule Molecular Inversion Probes allow cost-effective targeted sequencing of all genes and loci associated with macular diseases.. Invest. Ophthalmol. Vis. Sci. 2022;63(7):41.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Inherited retinal diseases are clinically and genetically heterogeneous. Amongst these are macular diseases (MDs) characterized by central vision loss. Inherited MDs (iMDs) are relatively rare and generally present at an early age, while age-related MD (AMD) is more prevalent and occurs later in life. The clinical and genetic heterogeneity in MDs makes a clear diagnosis challenging. The use of cost-effective single molecule Molecular Inversion Probes (smMIPs) to sequence ABCA4 in patients with Stargardt disease identified many coding and non-coding variants, but ~57% remained unsolved (Khan et al., 2020; PMID: 32307445). We hypothesized that the missing heritability may be revealed by smMIPs-based sequencing of all MD-associated genes.

Methods : We used 17,394 smMIPs designed by Molecular Loop Biosciences (USA) to sequence the coding regions of 105 iMD and AMD-associated genes, known pseudo-exons, and the mitochondrial genome. DNA libraries of 384 iMD samples previously investigated for variants in ABCA4, were pooled for sequencing on the NovaSeq 6000 platform. Variant filtering prioritized autosomal recessive variants with an allele frequency of ≤0.5% in population databases, and autosomal dominant variants with an allele frequency of ≤0.1%. Variants with a Franklin-ACMG classification of class 3-5 were selected.

Results : Across NovaSeq SP sequencing runs, an average nucleotide coverage of 680× was observed. Sequencing data for 104 iMD probands, who previously were shown not to carry two alleles in ABCA4, have been fully analysed. Across these 104 probands, 132 candidate variants in 40 genes, comprising two CNVs and 130 SNVs or indels, were identified. Of the SNVs or indels, 19 were class 5 variants (pathogenic), 21 class 4 variants (likely pathogenic) and 90 class 3 variants (VUS). Forty probands were considered to be genetically solved by disease causing variants in 24 genes, achieving a diagnostic yield of 38% within this ABCA4-pre-screened cohort.

Conclusions : Utilising smMIPs to target MD-associated genes and loci is a cost-effective approach. Further analysis will identify known and novel variants in iMD-associated genes, which will offer an accurate clinical diagnosis to patients and their families in addition to revealing new genetic associations for MDs.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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