A RIBOZYME WHICH DISCRIMINATES IN-VITRO BETWEEN PML RAR-ALPHA, THE T(15-17)-ASSOCIATED FUSION RNA OF ACUTE PROMYELOCYTIC LEUKEMIA, AND PML AND RAR-ALPHA, THE TRANSCRIPTS FROM THE NONREARRANGED ALLELES/

Acute promyelocytic leukemia (FAB M3) is distinguished by the presence of the t(15;17) and clinical response to all-trans retinoic acid (RA) treatment. Acute promyelocytic leukemia is associated with a chromoso mal translocation which results in the fusion of genes encoding a puta tive transcription factor (PML) and the retinoic acid receptor alpha ( RAR alpha). It is suggested that the PML/RAR alpha fusion protein func tions as an inhibitor of myeloid differentiation. The potential use of ribozymes as therapeutic agents has been investigated in the present study, Hammerhead ribozymes, which by hybridizing to both PML and RAR alpha sequences discriminate between the fusion transcript and the nor mal transcripts from the nonrearranged alleles, were designed and synt hesized, Two hammerhead cleavage sites were targeted: site 1, an AUU l ocated 4 nucleotides 3' to the fusion junction; and site 2, a UUC loca ted 26 nucleotides 3' to the junction, Both sites are located in the R AR alpha portion of the fusion transcript. Using a full-length PML/RAR alpha RNA or an RNA corresponding to 788 nucleotides of the PML/RAR a lpha mRNA and a full-length RAR alpha RNA or an RNA corresponding to 9 60 nucleotides of the RAR alpha mRNA as model substrates, the catalyti c behavior of several ribozymes was studied. A modified hammerhead dir ected against site 2 displayed the highest degree of selectivity for P ML/RAR alpha. It is hypothesized that ribozyme-mediated inactivation o f PML/RAR alpha provides a new approach to study the role of PML/RAR a lpha in the deregulated growth and RA response of acute promyelocytic leukemia.