Résumé :
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Spinal Muscular Atrophy (SMA) is a group of autosomal recessive neuromuscular diseases, characterized by progressive symmetrical muscle weakness with muscle atrophy. Deletion of the Survival Motor Neuron1 (SMN1) gene causes degeneration of anterior horn cells of the spinal cord. Although SMN2, a centromeric copy of SMN1, is present, a single base difference in exon 7 of SMN2 leads to exclusion of exon 7 which, in turn, disrupts the reading frame and dramatically reduces the amount of functional protein. Antisense Oligomers (AOs) have been widely used in gene inactivation applications. Oligomer action is determined by the base composition, chemistry, and nature of the backbone, thereby allowing modification of gene expression at different stages of pre-mRNA processing. AOs can modify pre-mRNA splicing by masking exonic sequence enhancer motifs (ESEs), involved in exon definition and splicing. ESEs, which are well-defined as positive regulatory elements appear crucial to splicing, as many mutations disrupting ESEs cause exon skipping. On the contrary, splice silencing motifs are less well-characterized. To induce exon inclusion, we designed an array of AOs to identify and suppress silencing splice motifs. The AOs were evaluated for exon inclusion efficiency in fibroblast cells derived from a SMA patient. Combinations of AOs were employed to assess interplay between different targeted motifs involving in pre-mRNA splicing. Several potential silencing motifs were identified as determined by RNA and protein studies. This preliminary data will be used to design additional AOs for efficient exon 7 inclusion.
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