New Therapeutic Approaches and Prognostic Assays for Breast Cancer: Padiolabeled Ligands and Antibodies and Quantitative PCR
Abstract
The presence of estrogen receptor (ER) alpha in breast tumors is used as a marker to predict prognosis and treat patients with anti-estrogen therapies. The rate of response to anti-estrogen therapies is about 50%. Recent studies have shown that tumor cells express several forms of ER suggesting that the composition of various isoforms in the tumor cells may correlate with response/resistance to anti-estrogen therapies. Currently, the tumor tissues are diagnosed for the presence of ER alpha by immunohistochemistry The problem is that this method is not sensitive to detect small amounts of ER protein and cannot distinguish and quantify various ER isoforms. For these reasons, there is an urgent need to develop a highly sensitive assay that can rapidly quantify various ER isoforms. We proposed to develop PCR based molecular assays that can quantify ER mRNAs. We used reverse transcription template competition approach to devise an assay and showed that it could be applied to quantify the major tumor ER mRNA, the alpha isoform, in the tumor tissues. Since only a few of the alternatively spliced ER mRNAs could be identified by co-amplification with the wild type ER, we developed a Splice Targeted Primer Approach' to specifically amplify the splice variants with out amplifying the wild type. Using this novel approach, we identified twenty different alternatively spliced ER mRNAs in breast cancer cell lines and tumors many of which were not known previously. We have also developed methodologies that can quantify the alternatively spliced ER alpha mRNAs as separate gene populations using the splice targeted primers and template competition PCR. These methods will be highly valuable in establishing a correlation between the expression of various ERs in a tumor with response/resistance to anti-estrogen therapies.
Document Details
- Document Type
- Technical Report
- Publication Date
- Oct 01, 1999
- Accession Number
- ADA381559
Entities
People
- Indra Poola
Organizations
- Howard University