Myotonic dystrophy type 1 (DM1) is an inherited neuromuscular disorder, which affects approximately 1 in 8000 people, and for which there are no treatments. The available evidence supports a toxic RNA gain of function mechanism in which CUG repeat RNA sequesters MBNL1, a regulator of alternative splicing, and indirectly increases levels of the protein CUGBP. In the parent proposal we are using rational design, computational methods, and high throughput screening to identify molecules that bind to CUG repeat RNA and inhibit complex formation with MBNL1 protein in order to validate CUG repeat RNA as a target for the treatment of DM1, and to identify compounds for optimization and evaluation in pre-clinical models of DM1. Using rational design, we have identified a triaminotriazine-acridine conjugate that binds selectively to CUG repeat RNA and inhibits binding of MBNL1 protein. In this supplement, we propose to establish a new collaboration with the Berglund laboratory at the University of Oregon to use X-ray crystallography to determine the structure of the triaminotriazine-acridine ligand bound to CUG repeat RNA. Berglund is an established RNA crystallographer who solved the first crystal structure of CUG repeats and has obtained new structures of CUG repeats that will be used to obtain structures of the complexes with triaminotriazine-acridine ligands. We view the proposed project as the establishment of a long-term collaboration with the Berglund group. None of the PI's on the parent proposal is experienced with X-ray crystallography, and therefore, the proposed work could not proceed without this collaboration. Structural characterization of ligand-RNA complexes will be essential to rationally develop the next generation triaminotriazine-acridine ligands or the next generation of compounds identified by high throughput screening or computational methods. There are relatively few ligand-RNA complexes that have been structurally characterized, and therefore, there is only a limited predictive knowledge of RNA-small molecule complex structures. Structural characterization will allow existing individual interactions to direct the construction of targeted libraries of compounds biased towards improved recognition. The addition of the Berglund group to the parent project forms a team with considerable expertise in organic synthesis, computational docking, high throughput screening, structure determination, cellular assays, and with the existing collaboration with the Thornton group at the University of Rochester, evaluation in animal models. Although the project is ambitious, we believe it can be accomplished in one year because the ligands are straightforward to synthesize, and the Berglund group has crystals of the CUG repeat RNA. The innovation and impact of this proposal lies in the selection of the RNA target involved in a disease for which there is no treatment, the multipronged approach to discover new classes of ligands, and with this supplement, the use of structural information to guide and inform the rational design and selection of specific, functional RNA binding ligands.
Myotonic dystrophy (DM1) afflicts approximately 1 in 8000 adults and is characterized by progressive muscular weakness, cardiac defect, cataracts, and other neuromuscular problems. We propose to develop a novel treatment strategy for DM1 that involves using small molecules to directly target the toxic RNA, the RNA that prevents important proteins from performing their normal functions. We are establishing a new collaboration to use X-ray crystallography to guide the design of the small molecule ligands. This approach may serve as a prototype for the treatment of other diseases caused by toxic RNAs.
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