Repression of human thymidylate synthase mRNA translation by antisense 2'-O-methyl oligoribonucleotides

John C. Schmitz, Sudhir Agrawal, Edward Chu

Research output: Contribution to journalArticlepeer-review

7 Scopus citations

Abstract

Previous studies have shown that translation of thymidylate synthase (TS) mRNA is controlled by its own protein end product TS in a negative autoregulatory manner. Disruption of this process results in increased synthesis of TS and may be associated with the development of cellular drug resistance to TS-directed anticancer agents. As one strategy to inhibit TS expression, we have designed antisense RNA oligoribonucleotides (ORNs) that directly target the 5'-upstream binding site (nt 80-109) of TS mRNA, a critical cis-acting regulatory element. ORNs were analyzed for their ability to specifically inhibit translation of human TS mRNA in an in vitro rabbit reticulocyte lysate translation system. Native 2'-hydroxyl(OH) ORNs inhibited TS mRNA translation in a dose-dependent manner but did not repress translation of control mRNAs, including p53 or Escherichia coli TS. A control sense 2'-OH ORN was unable to repress translation of either human TS mRNA, or control mRNAs. Modified antisense ORNs with 2'-O-methyl phosphodiester or 2'-O-methyl phosphorothioate backbones (or both) repressed human TS mRNA translation in a dose-dependent manner, and they were both more effective than the respective 2'-OH ORN. However, nonspecific effects on mRNA translation were observed with the 2'-O-methyl phosphorothioate ORN. In vitro translation experiments revealed that in the presence of antisense ORNs, the target TS mRNA remained intact. These findings demonstrate that antisense ORNs targeted at the 5'-upstream cis-acting element represent effective inhibitors of TS mRNA translation.

Original languageEnglish (US)
Pages (from-to)371-378
Number of pages8
JournalAntisense and Nucleic Acid Drug Development
Volume8
Issue number5
DOIs
StatePublished - 1998
Externally publishedYes

ASJC Scopus subject areas

  • Genetics
  • Pharmacology

Fingerprint Dive into the research topics of 'Repression of human thymidylate synthase mRNA translation by antisense 2'-O-methyl oligoribonucleotides'. Together they form a unique fingerprint.

Cite this