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.
|Number of pages||10|
|Journal||Genetics in medicine : official journal of the American College of Medical Genetics|
|Publication status||Published - Aug 2019|
- ATP-Binding Cassette Transporters/genetics
- HEK293 Cells
- Middle Aged
- Oligonucleotides, Antisense/genetics
- Polymorphism, Single Nucleotide/genetics
- Protein Isoforms/genetics
- RNA Splicing/genetics
- Stargardt Disease/genetics
- Young Adult