Potent Growth Inhibition of Leukemic Cells by Novel Ribbon-type Antisense Oligonucleotides to c-myb1

Abstract

We studied the effects of antisense oligonucleotides (AS oligos) with a novel structure. The AS oligos were covalently closed to avoid exonuclease activities by enzymatic ligation of two identical molecules. The AS oligos of a ribbon type (RiAS oligos) consist of two loops containing multiple antisense sequences and a stem connecting the two loops. Three antisense sequences targeting different binding sites were placed in a loop that was designed to form a minimal secondary structure by itself. RiAS oligos were found to be stable because they largely preserved their structural integrity after 24 h incubation in the presence of either exonuclease III or serums. When a human promyelocytic cell line, HL-60, was treated with RiAS oligos to c-myb, c-myb expression was effectively ablated. Cell growth was inhibited by >90% determined by both the 3-[4,5-dimethythiazol- 2-yl]-2,5-diphenyltetrazolium bromide assay and [3H]thymidine incorporation. Further, when the leukemic cell line K562 was treated with c-myb RiAS oligos, colony formation on soft agarose was reduced by 92 6 2%. These results suggest that RiAS oligos may be employed for developing molecular antisense drugs as well as for the functional study of a gene.