Regulation of EphA2 and focal adhesion kinase in breast cancer
Abstract
Changes in cell adhesion are required for cancer cells to metastasize and sites of cell adhesion are the primary location of tyrosine phosphorylated proteins. Identification and characterization of tyrosine phosphorylated proteins that localize to sites of cell adhesion might reveal new diagnostic and therapeutic targets. Two breast cancer cell line models were used to examine the regulation of EphA2 and the Focal Adhesion Kinase (FAK) by cell adhesion. Using a novel monoclonal antibody screen, EphA2 was identified as an adhesion-regulated, phosphorylated protein. EphA2 expression, tyrosine phosphorylation and localization, but not enyzmatic activity, distinguished cells derived from non-transformed mammary tissue, poorly invasive breast cancer, and metastatic breast cancer. Therefore, EphA2 may be useful as a diagnostic target. EphA2 expression is regulated by three factors known to play a role in breast cancer, as estrogen and Myc repressed EphA2 expression and Ras induced EphA2 expression. EphA2 phosphorylation and localization is regulated by E-cadherin, a cell-cell adhesion protein and known tumor suppressor. In non-transformed epithelial cells, EphA2 co-localized with E-cadherin. While disruption of E-cadherin mediated adhesion inhibited EphA2 tyrosine phosphorylation and cell-cell localization, re-expression of E-cadherin in metastatic breast cancer cells restored this phenotype. Alteration in EphA2 function may directly promote an invasive phenotype, as restoration of EphA2 phosphorylation in metastatic cells inhibited cell proliferation. Therefore, EphA2 may be useful as a therapeutic target. Cell-extracellular matrix (ECM) adhesion is essential for cancer cells to migrate. Characterization of the cell line models used in this study revealed that cells derived from poorly invasive breast cancer had the greatest cell-ECM adhesion and large focal adhesions. These changes in cell adhesion correlated with decreased FAK expression, but increased phosphorylation per FAK molecule. Alteration of cell-ECM adhesion by suspension and matrix binding assays demonstrated FAK expression and phosphorylation was adhesion dependent. Low FAK expression in tumor cells thus reflects the extent of cell adhesion rather than invasiveness directly. This may indicate a susceptibility to FAK directed anti-metastatic therapeutics.
Degree
Ph.D.
Advisors
Kinch, Purdue University.
Subject Area
Cellular biology|Oncology
Off-Campus Purdue Users:
To access this dissertation, please log in to our
proxy server.