Training in Support of research Project Entitled 'Tumor Cell De-Adhesion by Aberrant, Single Subunit Integrins'

摘要

I hypothesize that there are substantial differences in the plasma membrane complement of metastatic vs non- metastatic tumor cells. To test my hypothesis, I prepared plasma membrane proteins from three breast carcinoma cell lines (MDA-MB-435, MDA-MB-23 1 and MCF-7) of differing degrees of invasiveness, and generated 2D polypeptide expression maps. Computer analysis of the 2D polypeptide patterns showed that the two non-metastatic cell lines (MCF-7 and MDA-MB-23 1) displayed 56% similarity in their proteins, whereas MCF-7 and the highly metastatic cell line, MDA-MB-435, displayed only 42% similarity in their proteins. Further analysis showed 81 polypeptide spots to be unique to the 2D gel of the MDA-MB-43 5 cell line. Five of these proteins were identified using techniques of mass spectrometry and database searching. As an additional method of identifying plasma membrane proteins with relevance to tumor progression, I applied the technique of antibody library phage display. I created scFv antibody libraries to the plasma membrane proteins of MCF-7 and MDA-MB-43 5 cells and enriched these libraries for antibodies to cell-surface proteins. By flow cytometry, I identified an individual phage clone that displays an antibody that recognizes a unique protein on the surface of the metastatic cells. The identification of proteins that are unique to metastatic cells will increase our knowledge of the molecular mechanisms that govern tumor progression.