AON-mediated Exon Skipping Restores Ciliation in Fibroblasts Harboring the Common Leber Congenital Amaurosis CEP290 Mutation

Abstract

Leber congenital amaurosis (LCA) is a severe hereditary retinal dystrophy responsible for congenital or early-onset blindness. The most common disease-causing mutation (>10%) is located deep in intron 26 of the CEP290 gene (c.2991+1655A>G). It creates a strong splice donor site that leads to insertion of a cryptic exon encoding a premature stop codon. In the present study, we show that the use of antisense oligonucleotides (AONs) allow an efficient skipping of the mutant cryptic exon and the restoration of ciliation in fibroblasts of affected patients. These data support the feasibility of an AON-mediated exon skipping strategy to correct the aberrant splicing.

Antisense Oligonucleotide Therapy for Inherited Retinal Dystrophies

“…An ideal candidate for AON-based therapy is a recurrent intronic mutation in CEP290 (c.2991 + 1655A > G) that is causative for up to 15 % of all LCA cases in the US and several European countries (den Hollander et al 2006;Perrault et al 2007;Stone 2007;Coppieters et al 2010). This mutation activates a cryptic splice donor site that results in the insertion of a pseudo-exon with a premature termination codon to approximately 50-75 % of the CEP290 transcripts (den Hollander et al 2006;Gerard et al 2012). We have shown that, in lymphoblastoid and fibroblast cells from LCA patients with a homozygous intronic CEP290 mutation, administration of AONs targeting the pseudo-exon fully restores normal CEP290 pre-mRNA splicing (Collin et al 2012;Gerard et al 2012) (Fig.…”

Formulation of pH responsive peptides as inhalable dry powders for pulmonary delivery of nucleic acids

“…Our previous study has demonstrated that the pH responsive peptides are capable of mediating both highly efficient DNA or siRNA transfection in mammalian cell lines [15] (Abbate et al, unpublished). Others have shown that LAH4 peptides can effectively deliver nucleic acids to patient derived primary fibroblasts [18] and facilitate intracellular delivery in vivo by way of subcutaneous injection of protein-based vaccines adjuvanted with Toll-like receptor 9 agonist CpG oligonucleotide (CpG) to generate enhanced CD8+ T cell immune responses and antitumor effects [19]. With proven efficacy in delivery to a variety of cultured, primary cells and tumour cells together with the first example of in vivo tolerance and efficacy, we were interested to develop a pH responsive peptide-based formulation that is suitable for effective nucleic acid delivery by pulmonary administration.…”

Stem cells with a view: a look inside a retinal ciliopathy

“…In fact, AON-mediated correction of the c.2991 + 1665A > G CEP290 mutation in patientderived fibroblasts was demonstrated in two independent studies (19,20). The corrected gene exhibited improved transcription of the wild type transcript and restoration of cilia formation in the patient fibroblasts.…”