Differential Phosphoprotein Profiling of Tamoxifen Response
Abstract
Tamoxifen revolutionized breast cancer treatment when it came into use some three decades ago. In estrogen receptor(ER)-? positive breast cancer cells, tamoxifen blocks cancer growth by competing for binding to ER and cuts recurrence risk in half. However, some advanced breast cancers that initially respond well to tamoxifen eventually become resistant. Several mechanisms of resistance have been hypothesized, including crosstalk between ER and growth factor receptor tyrosine kinase pathways 1. Several studies suggest that overexpression of HER2, and high levels of phosphorylated Akt or ERK, contribute to tamoxifen resistance. By cataloging global phosphorylation events in response to tamoxifen treatment in tamoxifen sensitive and resistant cells we will provide better understanding of the mechanism of acquired resistance and/or identify novel biomarkers for tamoxifen response. We have developed a method for comparison of global phosphoprotein profiles. Our methodology involves stable isotope labeling 2, a phosphoprotein affinity step, 1-D SDS-PAGE and LC-MS/MS 3. I have shown differential phosphoprotein patterns in MCF7/Chicago (tamoxifen sensitive) and MCF7/HER2 (tamoxifen resistant) cells as a result of tamoxifen treatment. I present preliminary data towards phosphoprotein profiling of tamoxifen response in the cytoplasmic fraction of MCF7/Chicago cells. Over 1250 proteins were identified using an Orbitrap (Thermo) including over 40 kinases and 20 phosphatases. A subset of the proteins is isotopically labeled and quantitative revealed that a majority of the proteins did not change in abundance, as expected. However, about 60 proteins were identified with Xpress ratio less than 0.6 and about 28 proteins with ratio larger than 1.66. Manual analysis is underway to confirm the protein abundance ratios.
Document Details
- Document Type
- Technical Report
- Publication Date
- Aug 01, 2008
- Accession Number
- ADA502498
Entities
People
- Kolbrun Kristjansdottir
Organizations
- University of Chicago