SPLICEOSOME TRANS-SPLICING TO REPAIR CF MRNA

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The ultimate objective of this proposal is to develop a new, general mechanism to reprogram any targeted gene at the pre-messenger RNA level. Specifically, this proposal will develop the technology to produce a gene therapy for the treatment of Cystic Fibrosis. In Phase I, we will produce a series of pre-therapeutic RNA molecules (PTMs) that are capable of promoting spliceosome mediated RNA trans-splicing reactions with mutant Cystic Fibrosis Transmembrane Regulator (CFTR) pre- mRNA, repairing mutant mRNA in cultured cells from CF patients. The immediate goals are: 1) to develop one or more PTMs that can repair mutant CFTR mRNA; and 2) express corrected CFTR protein and restore chloride channel function in cultured cells. Currently, we have one PTM that is capable of repairing CFTR exon 10, which includes the most prevalent mutation, delta F508. Modifications of this PTM, shorted PTMs, and PTMs targeting different CFTR regions will be produced and functionally tested under this proposal. Other genetic diseases may also be amenable to trans-splicing RNA repair. Trans-splicing PTMs could improve gene therapies by conferring intra-cellular specificity, decreasing the size of the delivered gene, and acquiring the expression pattern of the endogenous target gene. PROPOSED COMMERCIAL APPLICATIONS: Development of RNA molecules capable of repairing mutant CFTR by trans-splicing could lead to a therapy for Cystic Fibrosis that could slow or halt disease progression. There is a clinical need to serve and treat this market of 30,000 affected individuals in the U.S. Additionally, the development of effective spliceosome mediated RNA trans-splicing technology would be of utility in many gene transfer applications including the treatment of other genetic diseases, infection by splicing viruses (HIV, EV, papilloma, etc), and cancer.