Marfan syndrome (MFS) is a dominant connective-tissue disorder caused by fibrillin-1 (FBN1) mutations. Fibrillin-1, is a large glycoprotein that aggregates into multimers to form the backbone of microfibrils, providing structural and regulatory support to connective tissues. MFS shows variable age of onset and is progressive in nature, with major clinical manifestations in the skeletal, ocular and cardiovascular systems. The aetiology of MFS is not fully understood, however, mutations in FBN1 are thought to result in heterogeneous fibrillin-1 monomers that are unable to form multimers, leading to reduction in microfibril abundance. To date, over 1,800 unique mutations are reported in FBN1, including a c.6354C>T mutation resulting in exon 52 skipping from ~half the processed mRNA transcripts. This study demonstrates a potential therapeutic strategy for MFS using antisense oligonucleotides to alter the exon structure, of both the normal and disease causing mRNA transcripts, to re-establish the periodicity of fibrillin-1. We hypothesise that the resulting modified fibrillin-1 monomers will correctly form multimers, increasing microfibril abundance and reducing disease severity.
Antisense oligonucleotides, designed to target regulatory splicing motifs within FBN1 exon 52, were screened in normal and patient fibroblasts, to assess induced exon 52 skipping from the mRNA transcripts. Treated cells were also immunostained using an anti-fibrillin-1 antibody to reveal the abundance and morphology of microfibrils. The observed exon 52 skipping was dose dependant, with up to 99% of transcripts in patient fibroblasts missing exon 52. A corresponding increase in microfibril abundance was observed in treated, compared to untreated patient cells
The use of antisense oligonucleotides to induce targeted alternative splicing has garnered attention in recent years, particularly for the treatment of Duchenne muscular dystrophy. We believe this technique is applicable to MFS, with preliminary in vitro data supporting the hypothesis that inducing homogeneity between fibrillin-1 monomers is a potential therapeutic option.