Specific down-regulation of HER-2/neu mediated by a chimeric U6 hammerhead ribozyme results in growth inhibition of human ovarian carcinoma

Vivian W.Y. Lui, Yukai He, Kirti Goyaland, Leaf Huang

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

The U6 expression system was explored for efficient expression of a ribozyme against the human proto-oncogene c-neu. A hammerhead ribozyme (neuRz) and the control mutant ribozyme (MRz) were targeted to cleave c-neu mRNA at the tyrosine kinase domain. In vitro cleavage showed that neuRz was very active while MRz was not. Near-maximal target cleavage observed at a low ribozyme:target ratio (0.1) suggests that neuRz has good activity and turnover capability under physiological conditions, i.e., <5 mM MgCI2 and 37°C. Chimeric U6 ribozyme was expressed at about 5 × 106 copies/cell at 48 h in the ovarian carcinoma cell line SKOV-3.ip1. Partial down-regulation of c-neu mRNA and protein was observed in a dose-dependent manner in cells transiently transfected with U6neuRz- but not with MRz-containing plasmid. Sorted transient transfectants demonstrated dramatic growth inhibition with the neuRz-expressing cells. Our results demonstrate that the U6 expression system is very efficient and suitable for the expression of a hammerhead ribozyme. Moreover, nonviral delivery of the neuRz-expressing plasmid resulted in specific down-regulation of c-neu and, subsequently, growth inhibition of ovarian cancer cells overexpressing c-neu.

Original languageEnglish (US)
Pages (from-to)169-177
Number of pages9
JournalMolecular Therapy
Volume3
Issue number2
DOIs
StatePublished - Jan 1 2001
Externally publishedYes

Keywords

  • Gene therapy
  • HER-2/neu ribozyme
  • Ovarian cancer growth inhibition
  • U6 RNA

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Pharmacology
  • Drug Discovery

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