A Cell-Based Double Reporter Gene Splicing Assay for Therapeutic Screening in Myotonic Dystrophy

Abstract The study has developed a model splicing construct assay system based on splicing misregulation, one of the major molecular features associated with myotonic dystrophy. The splicing construct assay has double reporters for intron 2 splicing in chloride channel (CLCN1). The CLCN1 transgene splicing construct assay was used to transfect wild type and DM fibroblast cell lines and the clones generated showed that it enabled quantification of splicing efficiency in transgene construct. Validation of the DM fibroblasts containing transgene splicing construct was performed by differentiating the DM fibroblasts into myoblasts which exhibited a switch in CLCN1 splicing construct which was consistent with that associated with myotonic dystrophy (DM) condition. The myoblast derived from fibroblasts cell-based gene-splicing assay was subsequently applied in therapeutic screening in small throughput screens of 113 compounds which identified Protein Kinase C inhibitors- hypericin and Ro-31-8220 as potential therapeutic agents. The CLCN1 gene-splicing assay is a good model system for application in therapeutic screening in myotonic dystrophy because its double reporters facilitated quantification of effect putative drug on correction of misregulated splicing.