SELECTIVE CLEAVAGE OF BCR-ABL CHIMERIC RNAS BY A RIBOZYME TARGETED TONONCONTIGUOUS SEQUENCES

Conventionally designed ribozymes may be unable to cleave RNA at sites which are inaccessible due to secondary structure. In addition, it ma y also be difficult to specifically target a conventionally designed r ibozyme to some chimeric RNA molecules. Novel approaches for ribozyme targeting were developed by using the L6 bcr-abl fusion RNA as a model . Using one approach, we successfully directed ribozyme nucleation to a site on the bcr-abl RNA that is distant from the GUA cleavage site. These ribozymes bound to the L6 substrate RNA via an anchor sequence t hat was complementary to bcr sequences. The anchor was necessary for e fficient cleavage as the anchor minus ribozyme, a conventionally desig ned ribozyme, was inefficient at catalyzing cleavage at this same site . The effect of anchor sequences on catalytic rates was determined for two of these ribozymes. Ribozymes generated by a second approach were designed to cleave at a CUU site in proximity to the bcr-abl junction . Both approaches have led to the development of a series of ribozymes specific for both the L6 and K28 bcr-abl chimeric RNAs, but not norma l abl or bcr RNAs. The specificity of the ribozyme correlated in part with the ability of the ribozyme to bind substrate as demonstrated by gel shift analyses. Secondary structure predictions for the RNA substr ate support the experimental results and may prove useful as a theoret ical basis for the design of ribozymes.