Type VII collagen re-expression and anchoring fibrils formation after systemic delivery of an antisense oligonucleotide inducing COL7A1 exon 73 skipping for RDEB.

Summary: Abstract Body: Skipping of the frequently mutated COL7A1 exon 73 using antisense oligonucleotide (ASO) is a promising approach to treat recessive dystrophic epidermolysis bullosa (RDEB), a rare genodermatosis caused by loss-of-function variants in COL7A1 encoding type VII collagen (C7). RDEB is characterized by severe blistering of the skin and mucosae, including oesophageal strictures and corneal blisters. We have developed a palmitoylated tricyclo-DNA ASO which induces efficient exon 73 skipping and C7 re-expression in vitro after transfection in primary RDEB keratinocytes and fibroblasts. We grafted reconstructed skin from C7-deficient RDEB patient cells onto an immunodeficient murine model (NMRI-Foxn1nu/nu) and demonstrated restoration of C7 expression and formation of mature anchoring fibrils (AF) at the dermal-epidermal junction up to one month after the last injection of either two subcutaneous injections of 500 µg or 1 mg of the ASO beneath the graft, or 4 or 8 weekly intravenous injections at 50mg/kg. This ASO was also injected intravenously in a transgenic murine model (mCol7a1-/-; TghCOL7A1) carrying the entire human COL7A1 genomic locus. Transcript analyses of disease relevant tissues (skin, eyes, and oesophagus) revealed variable levels of exon 73 skipping in vivo, consistent with ASO biodistribution. No adverse reaction was observed and in vitro pre-toxicology studies showed that the ASO does not form homodimers nor activate complement. Overall, our data indicate that systemic administration of palmitoylated tricyclo-DNA ASO restores C7 expression and AF formation by skipping exon 73 of COL7A1 pre-mRNA and holds therapeutic potential for RDEB patients. Guillaume Mondon¹, Vincent Nguyen¹, Daniela Vieira², Giovanna Zambruno³, Luis Garcia², Alain Hovnanian¹, Matthias Titeux¹ 1. Paris Cité University - INSERM U1163 - Imagine Institute, Paris, France. 2. INSERM U1179 - Versailles Saint-Quentin-en-Yvelines University, Montigny-le-Bretonneux, France. 3. Bambino Gesù Children’s Hospital, Rome, Italy. Genetic Disease, Gene Regulation, Gene Therapy & Epigenetics