Circular antisense oligonucleotides inhibit growth of chronic myeloid leukemia cells

Background: Antisense represents a conceptually powerful method for regulat ing gene expression. However, antisense oligonucleotides developed to date manifest two serious limitations-nuclease susceptibility and nonspecific hy bridization. Circular oligonucleotides may be superior to conventional line ar oligonucleotides in both respects. First, circular agents, having no end s, are exonuclease-resistant. Second, they bind to complementary strands of RNA and DNA with a higher affinity than corresponding linear agents. Methods and Results: We assessed the activity of circular phosphodiester de oxynucleotides using chronic myeloid cell lines by targeting polypurine seq uences. To represent cells having a bcr3/abl2-type junction, we used K562 c ells. A circle targeting a bcr polypurine sequence 385 nucleotides 5' to th e junction decreased the cell number by day 5 with an IC50 of 9 mu M. To re present cells having a bcr2/abl2-type junction, we used BV173 cells. A circ le targeting the bcr-abl junction itself decreased the cell number by day 7 with an IC50 of 8 mu M. Control oligonucleotides, whether the same sequenc e uncircularized or circles with the same nucleotide composition but in scr ambled sequence, had little effect. Unlike linear agents, circles were stab le when incubated in 10% serum. The amount of ba-abl protein detected by We stern blotting using a specific anti-ba-abl antibody at 24 hr in antisense- treated BV173 cells was only 10% of that of cells treated with control circ les, which demonstrates an antisense mechanism of action. Conclusions: Circular oligodeoxyribonucleotides (1) inhibit the accumulatio n of CML cells, (2) decrease the amount of bcr-abl protein per cell, (3) ha ve sequence-selective activity, and (4) are more active than linear oligonu cleotides containing only the base-pairing region.