Antibody Probes to Estrogen Receptor-alpha Transcript-Specific Upstream Peptides: Alternates ER-alpha Promoter Use and Breast Cancer Etiology/Outcome
Abstract
Estrogen Receptor alpha (ER)expression correlates with a reduced incidence of breast cancer recurrence following resection of tumors. ER-protein in breast tumors increases with patient age. The project developed antibody reagents to probe expression of ER from alternate promoters taking advantage of short peptides encoded by upstream open reading frames (uORFs) in regions of the ER mRNAs that are promoter specific and not shared. We hoped that the short transcript specific peptides would act as Biomarkers' of promoter use. We developed polyclonal and monoclonal reagents effective in detecting the uORF peptides of the proximal and distal ER promoter transcripts. Antisera were validated using peptides linked to Biacore chips and in western blots using fusion of the peptides to larger proteins. The small size (2000 Da) precluded direct detection in immunoblots. However, the project stalled at the point of detecting the peptide in actual tumor samples by immunochemistry. The major problem has been that the antisera cross react to other proteins giving a non-specific background. This probably relates to small sized of the peptides - providing limited numbers of antigenic features. It may be possible to use the antisera as part of the search for the peptides in cells using mass spectroscopy.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 01, 2005
- Accession Number
- ADA438213
Entities
People
- Brian T. Pentecost
Organizations
- Health Research, Incorporated