Inhibition of DNA polymerase α expression and cell growth, a possible triple helix mechanism

The antiproliferative effects mediated by a 14-mer homopyrimidine oligonucleotide (5' CTTTCTCTTTTCTC3' designed to form DNA triplex with a purine region of the DNA polymerase α promoter, were evaluated on the human breast cancer cell line MDA-MB 231. In order to stabilize the triplex complex under physiologic conditions, replacement of cytosines by methylcytosines in the oligomer sequence was carried out. Band-shift analyses demonstrated a complete triplex formation between the radiolabeled target duplex DNA and the methylcytosine-modified oligomer at the concentration of 0.1 μM under physiologic pH and temperature. A single exposure of MDA-MB 231 cells to 0.5 μM methylcytosine-modified oligonucleotide was able to markedly reduce the cell number and the percentage of cells in DNA synthesis up to 58% and 66%, respectively, compared with controls. Furthermore, a 48% reduction in the amount of the DNA polymerase a mRNA was reported after treatment with the oligomer. In conclusion, data from the present study demonstrate that an oligonucleotide to DNA polymerase α promoter, designed to form a triple helix with target double-stranded DNA, inhibits the expression of the reporter gene at the biologic and molecular levels, suggesting a possible triplex-mediated mechanism of action.