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. 1992 May;89(5):1523–1527. doi: 10.1172/JCI115744

c-myc antisense oligonucleotides inhibit the colony-forming capacity of Colo 320 colonic carcinoma cells.

J F Collins 1, P Herman 1, C Schuch 1, G C Bagby Jr 1
PMCID: PMC443024  PMID: 1569190

Abstract

Colo 320 cells are colonic carcinoma cells known to express abundant c-myc mRNA. Based on the response of several hematopoietic cell lines to chemical inducers of differentiation, we reasoned that such agents might have similar inductive activity in Colo 320 cells. Accordingly, we exposed Colo 320 cells to 5 mM sodium butyrate (NaBT) for 7 d. C-myc expression decreased threefold and self-replicative potential decreased (defined as a greater than 60% decrease in colony-forming capacity in soft agar that did not contain inducer). In an effort to demonstrate a direct cause and effect between myc expression and the colony-forming capacity of Colo 320 cells, we exposed these cells to a 15-base antisense c-myc oligonucleotide (complementary to the translation initiation region of exon II). Cells were also exposed to equimolar (20 microM) amounts of sense and missense oligonucleotides. Subsequently, cells were incubated at 10, 20, 30, and 40 microM antisense DNA for 16 h, then washed and plated in oligonucleotide-free agar medium. We demonstrated that: (a) the oligomers were stable in the extracellular medium and in the cell cytoplasm; (b) the uptake of the oligonucleotides was 0.7%; (c) sense and missense oligonucleotides had no effect on colony-forming capacity; and (d) the antisense c-myc oligonucleotide resulted in a 40-75% concentration-dependent decrease in colony-forming capacity. The specific inhibition of colony-forming capacity by antisense DNA suggests that the role of myc expression in Colo 320 cells is similar to its role in hematopoiesis, and that the failure to inhibit myc expression maintains colony-forming capacity. This system provides a new strategy for inducing differentiation and may provide further insight into the genetic factors that govern the process of colonic carcinogenesis.

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Selected References

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