title: Oligonucleotide N3'-->P5' phosphoramidates as efficient telomerase inhibitors.
abstract: Human telomerase is a unique reverse transcriptase that is expressed in multiple cancers, but not in the vast majority of normal cells. The enzyme is responsible for telomere protection and maintenance, and supports the proliferative immortality of cancer cells. Thus, it has been proposed that the specific inhibition of telomerase activity in tumors might have significant and beneficial therapeutic effects. To this goal we have designed, synthesized, and evaluated several oligonucleotide N3'-->P5' phosphoramidates as telomerase inhibitors. These oligonucleotides are complementary to the template region of the RNA domain of telomerase (hTR). The prepared compounds were evaluated in HME50-5E breast epithelial cells, where their effects on telomerase activity were determined using a cell-based telomerase (TRAP) assay at 24 as well as 72 h after exposure to compounds. The oligo-N3'-->P5' phosphoramidate inhibited telomerase activity in cells in the presence of the cellular up-take enhancer (FuGENE6) in a dose- and sequence-dependent manner, with IC(50) values of approximately 1 nM. Inhibition of telomerase activity by this compound without the lipid carrier was not efficient. However, the isosequential oligonucleotide N3'-->P5' thio-phosphoramidate was able to inhibit telomerase activity with or without lipid carriers at nM, or low-microM concentrations, respectively. This inhibition of telomerase activity in HME50-5E cells by the oligonucleotide thio-phosphoramidates was also sequence specific. Long-term treatment of the cells with 0.5 microM of FuGENE6 formulated 13-mer thio-phosphoramidates, fully complementary to hTR, resulted in gradual telomere shortening, followed by cellular senescence and apoptosis, as would be predicted for a telomerase inhibitor. The mismatched control compound had no effect on cell proliferation. The results suggest that the oligonucleotide N3'-->P5' phosphoramidates, and particularly thio-phosphoramidates, might be further developed as selective anti-telomerase reagents.
authors.affiliation: Department of Cell Biology, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, Texas, TX 75390-9039, USA.
authors.forename: Brittney-Shea
authors.initials: BS
authors.lastname: Herbert
authors.forename: Krisztina
authors.initials: K
authors.lastname: Pongracz
authors.forename: Jerry W
authors.initials: JW
authors.lastname: Shay
authors.forename: Sergei M
authors.initials: SM
authors.lastname: Gryaznov
authors.forename: Brittney
authors.initials: B
authors.lastname: Shea-Herbert
mesh_terms: D000577:Amides / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D002455:Cell Division; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000494:pharmacology; D005260:Female; D006801:Humans; D009841:Oligonucleotides / Q000737:chemistry / Q000494:pharmacology*; D010756:Phosphoric Acids / Q000494:pharmacology*; D019098:Telomerase / Q000037:antagonists & inhibitors* / Q000235:genetics; D016615:Telomere / Q000378:metabolism; D014407:Tumor Cells, Cultured
chemical_list: D000577:Amides; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D009841:Oligonucleotides; D010756:Phosphoric Acids; C011067:phosphoramidic acid; D019098:Telomerase