The therapeutic landscape for myotonic dystrophy type 1 (DM1) has advanced dramatically in the past 5 years, but conflicting observations about the metabolism of expanded CUG mRNA (transcription, export, turnover, and translation) present challenges for how to interpret results from clinical trials and direct future therapeutic strategies. Fully understanding these fundamental pathophysiological mechanisms in DM1 has been severely limited due to lack of an animal model containing repeat lengths reflective of human disease states and expressed with appropriate spatiotemporal dynamics. To address this problem, we have developed a new mouse model of DM1 (Dmpk CTGexp), which expresses >1700 CTG repeats within native mouse Dmpk. Heterozygous mice display disease features including nuclear foci, splicing defects, myotonia, and myopathy. Further, the expanded allele in this model is tagged with a single nucleotide polymorphism (SNP) for tracking mRNA accumulation, knockdown, and transport. By using this model, we will determine strengths and weaknesses of leading therapeutic strategies, helping to focus clinical efforts on those with greatest potential. Three therapeutic platforms – antisense oligonucleotides (ASOs), siRNAs, and phosphoramidite morpholino oligonucleotides (PMOs) – are in or are rapidly advancing toward early phase trials in DM1. Based on observations that mutant DMPK RNA sequesters MBNL proteins to form nuclear foci, causing extensive changes in the muscle transcriptome, these agents either degrade DMPK transcripts or release MBNL protein from nuclear foci by tiling the CUG repeat tract. While initial results are encouraging, there is vigorous debate surrounding the strengths and weaknesses of each strategy. In Aim 1, we will first clarify mechanisms regulating the life cycle of expanded Dmpk mRNA. We will study the effects of expanded CUG repeats on transcription, turnover, subcellular localization, and translation of host Dmpk mRNA. In Aim 2, we will compare three major therapeutic strategies: ASOs, siRNA, and PMOs. We will elucidate mechanisms, strengths, and limitations of each using validated skeletal muscle readouts. In Aim 3, we will evaluate multi-systemic treatments by testing a PMO conjugate and MyoAAV-miRNA, to test the hypothesis that such approaches can rescue splicing defects in skeletal, cardiac and/or smooth muscle.