Deep-intronic ABCA4 variants explain missing heritability in Stargardt disease and allow correction of splice defects by antisense oligonucleotides

Riccardo Sangermano, Alejandro Garanto, Mubeen Khan, Esmee H Runhart, Miriam Bauwens, Nathalie M Bax, L Ingeborgh van den Born, Muhammad Imran Khan, Stéphanie S Cornelis, Joke B G M Verheij, Jan-Willem R Pott, Alberta A H J Thiadens, Caroline C W Klaver, Bernard Puech, Isabelle Meunier, Sarah Naessens, Gavin Arno, Ana Fakin, Keren J Carss, F Lucy RaymondAndrew R Webster, Claire-Marie Dhaenens, Heidi Stöhr, Felix Grassmann, Bernhard H F Weber, Carel B Hoyng, Elfride De Baere, Silvia Albert, Rob W J Collin, Frans P M Cremers

Research output: Contribution to journalArticleResearchpeer-review


PURPOSE: Using exome sequencing, the underlying variants in many persons with autosomal recessive diseases remain undetected. We explored autosomal recessive Stargardt disease (STGD1) as a model to identify the missing heritability.

METHODS: Sequencing of ABCA4 was performed in 8 STGD1 cases with one variant and p.Asn1868Ile in trans, 25 cases with one variant, and 3 cases with no ABCA4 variant. The effect of intronic variants was analyzed using in vitro splice assays in HEK293T cells and patient-derived fibroblasts. Antisense oligonucleotides were used to correct splice defects.

RESULTS: In 24 of the probands (67%), one known and five novel deep-intronic variants were found. The five novel variants resulted in messenger RNA pseudoexon inclusions, due to strengthening of cryptic splice sites or by disrupting a splicing silencer motif. Variant c.769-784C>T showed partial insertion of a pseudoexon and was found in cis with c.5603A>T (p.Asn1868Ile), so its causal role could not be fully established. Variant c.4253+43G>A resulted in partial skipping of exon 28. Remarkably, antisense oligonucleotides targeting the aberrant splice processes resulted in (partial) correction of all splicing defects.

CONCLUSION: Our data demonstrate the importance of assessing noncoding variants in genetic diseases, and show the great potential of splice modulation therapy for deep-intronic variants.

Original languageEnglish
Pages (from-to)1751-1760
Number of pages10
JournalGenetics in medicine : official journal of the American College of Medical Genetics
Issue number8
Publication statusPublished - Aug 2019


  • ATP-Binding Cassette Transporters/genetics
  • Adolescent
  • Adult
  • Aged
  • Child
  • Exons/genetics
  • HEK293 Cells
  • Humans
  • Introns/genetics
  • Middle Aged
  • Mutation/genetics
  • Oligonucleotides, Antisense/genetics
  • Pedigree
  • Polymorphism, Single Nucleotide/genetics
  • Protein Isoforms/genetics
  • RNA Splicing/genetics
  • Stargardt Disease/genetics
  • Young Adult


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