Inactivation of FGF Receptors by Targeting Ribozymes Against FGFR mRNAs and their Effect on FGF Dependent In Vitro and In Vivo Breast Cancer Growth Phenotypes
Abstract
Acquired tamoxifen resistance is common in breast cancer patients with estrogen receptor positive (ER+) tumors. Growth factor signaling can provide ER+ breast cancer cells with alternative growth stimulus to that provided by activation of ER. In order to determine whether an individual FGF receptor (FGFR) or multiple receptors are responsible for conferring an alternate growth signaling pathway, we are using a ribozyme targeting strategy to selectively inactivate each of the receptors either singly or in combination. We have designed multiple ribozymes against each of the four receptors and tested their ability to cleave in vitro transcribed portions of FGFR RNA. These assays have indicated that at least one ribozyme against each receptor is able to efficiently cleave its target substrate in vitro. Next, we used transient transfection assays to determine the ability of each ribozyme to inhibit FGF-1 dependent phosphorylation of a cotransfected reporter encoding a Green Fluorescent Protein-MAPK fusion protein by Western blot analysis with phosphospecific antibodies. These studies indicate that ribozymes against individual FGFRs were not able to block phosphorylation of GFP-NAPK to the same extent as observed following cotransfection with a dominant negative FGFR.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 01, 2002
- Accession Number
- ADA405224
Entities
People
- Francis Kern
- Norman R. Estes
Organizations
- Southern Research