RNA interference as a therapeutic agent for neuromuscular disease

In many dominant genetic diseases the mutant gene that gives rise to the disease co-exists with a normal copy of the gene, frequently only differing from the normal copy by a single letter (nucleotide base) in its genetic code. Any method that was reliably able to discriminate the mutant copy of the gene from the normal copy (i.e. that was sensitive enough to discriminate as little as a single base difference), and destroy it, thereby leaving only normal copies of the gene present, would have great potential as a novel treatment strategy. Such a method now exists based on a powerful new technology called RNA interference (RNAi). We have studied dominant mutations in the muscle acetylcholine receptor (AChR) gene (that codes for an important neurotransmitter receptor essential for normal functioning of the nerve-muscle junction) that cause a disease called slow channel myasthenic syndrome. In this work we have demonstrated proof-of-concept for using RNAi to selectively silence expression of the mutant AChR genes. We now wish to develop further this very promising approach by; studying and optimizing the RNAi sequence and chemical structure for mutant gene silencing; testing these in muscle cell lines derived from patients with this disorder; investigating the parameters for effective and safe delivery of RNAi molecules to the nerve-muscle junction; and finally undertaking trials of this therapy in animals.

RNA interference (RNAi) is a powerful new method for gene silencing. We have demonstrated proof-of-principle for allele-specific gene silencing using RNAi, which offers a novel therapeutic approach for treating dominant genetic neurological disorders. We also have encouraging preliminary data on siRNA delivery to muscle in vivo. We therefore hypothesize that selective down regulation of mutant genes associated with neuromuscular junction (NMJ) disease is very likely to lessen the symptoms as well as slow the progression of the disease. This project will systematically investigate specific silencing of pathogenic mutant acetylcholine receptor (AChR) subunits that underlie the neuromuscular disorder, slow channel congenital myasthenic syndrome (SCCMS). We will identify highly potent RNAi molecules designed specifically to down regulate expression of genes associated with SCCMS; optimize the specific silencing of mutant AChR alleles in patient muscle cell lines; investigate and optimize RNAi delivery to the NMJ in vivo (testing both local and systemic RNAi delivery with chemically modified RNAi molecules and using viral expression and delivery); and conduct preclinical studies in a relevant animal model to evaluate its potential for correcting synaptic dysfunction. The ultimate goal of the research programme is to identify effective and safe candidate molecules and delivery systems for clinical application.