Splicing Modulation via Antisense Oligonucleotides in Recessive Dystrophic Epidermolysis Bullosa

Stefan Hainzl (First author), Lisa Trattner (First author), Bernadette Liemberger (Co-author), Johannes Bischof (Co-author), Thomas Kocher (Co-author), Michael Ablinger (Co-author), Alexander Nyström, Astrid Obermayer, Alfred Klausegger (Co-author), Christina Guttmann-Gruber (Co-author), Verena Wally (Co-author), Johann W Bauer (Co-author), Josefina Piñón Hofbauer (Last author), Ulrich Koller* (Last author)

*Corresponding author for this work

Research output: Contribution to journalOriginal Articlepeer-review

2 Citations (Web of Science)

Abstract

Antisense oligonucleotides (ASOs) represent an emerging therapeutic platform for targeting genetic diseases by influencing various aspects of (pre-)mRNA biology, such as splicing, stability, and translation. In this study, we investigated the potential of modulating the splicing pattern in recessive dystrophic epidermolysis bullosa (RDEB) patient cells carrying a frequent genomic variant (c.425A > G) that disrupts splicing in the COL7A1 gene by using short 2'-O-(2-Methoxyethyl) oligoribo-nucleotides (2'-MOE ASOs). COL7A1-encoded type VII collagen (C7) forms the anchoring fibrils within the skin that are essential for the attachment of the epidermis to the underlying dermis. As such, gene variants of COL7A1 leading to functionally impaired or absent C7 manifest in the form of extensive blistering and wounding. The severity of the disease pattern warrants the development of novel therapies for patients. The c.425A > G variant at the COL7A1 exon 3/intron 3 junction lowers the efficiency of splicing at this junction, resulting in non-functional C7 transcripts. However, we found that correct splicing still occurs, albeit at a very low level, highlighting an opportunity for intervention by modulating the splicing reaction. We therefore screened 2'-MOE ASOs that bind along the COL7A1 target region ranging from exon 3 to the intron 3/exon 4 junction for their ability to modulate splicing. We identified ASOs capable of increasing the relative levels of correctly spliced COL7A1 transcripts by RT-PCR, sqRT-PCR, and ddPCR. Furthermore, RDEB-derived skin equivalents treated with one of the most promising ASOs exhibited an increase in full-length C7 expression and its accurate deposition along the basement membrane zone (BMZ).

Original languageEnglish
Article number761
Number of pages17
JournalINTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Volume25
Issue number2
DOIs
Publication statusPublished - 7 Jan 2024

Keywords

  • Antisense oligonucleotides
  • Col7a1
  • Recessive dystrophic epidermolysis bullosa
  • Splicing modulation

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