RIBOZYME-MEDIATED TRANSSPLICING OF A TRINUCLEOTIDE REPEAT

Trinucleotide repeat expansions (TREs) are a recently described class of mutations characterized by a change in the size of the genomic frag ment due to amplification of the repeated unit. A number of diseases h ave been attributed to TRE, including Huntington disease and myotonic dystrophy (DM; refs 1-3), but attempts at genetic therapy have yet to prove successful. A potential therapeutic approach would be to repair the expanded repeat using the trans-splicing ability of group I intron ribozymes(4). We have used DM as a model to test this hypothesis. A g roup I intron ribozyme (DMPK-RZ1) was designed to modify the TRE at th e 3' end of the human myotonic dystrophy protein kinase (DMPK) transcr ipt(5-8). DMPK-RZ1 was shown to ligate a small DMPK mRNA fragment, con tained within the ribozyme, to a simple DMPK-target RNA in vitro. It a lso modified a larger target transcript, leading to replacement of twe lve repeats with five repeats, both in vitro and in mammalian cells. F inally, this ribozyme successfully replaced the 3' end of endogenous D MPK mRNA in fibroblasts with a different 3' region. Ribozyme-mediated RNA repair may thus form a novel therapeutic strategy for diseases ass ociated with repeat expansions.