Development of therapeutic agents for myotonic dystrophy type 1

Myotonic dystrophy type 1 (DM1) is a multisystemic neuromuscular disorder which is caused by a dominantly inherited (CTG·CAG) repeat expansion in the 3’-UTR of the Dystrophia Myotonica Protein Kinase (DMPK) gene on chromosome 19. The genetic basis of DM1 is that DMPK is transcribed into an mRNA transcript with an expanded CUG repeat, r(CUG)exp, which can form hairpin secondary structures. These hairpin structures sequester the muscleblind-like (MBNL) family of proteins. MBNL proteins are responsible for post-transcriptional splicing and polyadenylation site regulation. The sequestration of MBNL proteins prevents them from performing their proper function, leading to hundreds of splicing defects. DM1 therapeutic efforts have been focused on inhibiting the interaction between r(CUG)exp and MBNL1 proteins, because it is estimated that over 80% of DM1 splicing defects are related to MBNL1 sequestration. This dissertation focuses on the development of rationally designed small molecules that can function as potential therapeutic agents, including their synthesis and studies of their biological activity. The background of DM1 with a focus on its molecular mechanism and various therapeutic strategies are reviewed in Chapter 1. Chapter 2 reports a series of multitargeting oligomeric ligands that have been developed to serve as both drug molecules and as antisense delivery vehicles for DM1. As a result of multivalency, the longer oligomers bound to the target DNA and RNA with nanomolar affinities in vitro and exhibited improved disease outcomes in DM1 model cell culture and DM1 patient myotubes. In addition, the oligomer can form polyplexes with a locked nucleic acid (LNA) CAG16 gapmer, facilitating its cell uptake in both HeLa cells and DM1 patient myotubes, with a resulting decrease in the level of toxic r(CUG)exp significantly. RNA-seq analysis indicated that the oligomeric ligand and its polyplex can rescue DM1 associated splicing defects and gene expression changes transcriptome-wide in DM1 patient myotubes. Chapter 3 discusses a rationally designed minor groove binder for DM1 as a potential therapeutic agent. The ligand contains triaminotriazine as a selective recognition moiety for T-T and U-U mismatches, and a thiazole peptidomimetic linker as a minor groove binder. This ligand is shown to rescue DM1 related defects in vitro and in DM1 model cell culture, including decreasing toxic cellular r(CUG)exp level, foci dispersion, and Insulin Receptor (IR) pre-mRNA mis-splicing rescue.LimitedAuthor requested closed access (OA after 2yrs) in Vireo ETD syste