The Effect of Blood on Circulating Tumor Cells' Expression of Epithelial and Mesenchymal Cell Markers

Open Access
Author:
Clabbers, Andrea Laurie
Area of Honors:
Bioengineering
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Siyang Zheng, Thesis Supervisor
  • Sheereen Majd, Honors Advisor
  • William O Hancock, Faculty Reader
Keywords:
  • Circulating Tumor Cells
  • EpCAM
  • Epithelial to Mesenchymal Transition
Abstract:
This research aims to examine the variation in expression of the biological markers EpCAM, vimentin, and cytokeratin on the surface of circulating tumor cells (CTC) depending on the cells’ time in blood. CTC detection, enumeration, and characterization offer potential prognostic information in a much less invasive form than current bone marrow biopsies. CTC that are especially of interest are ones that are undergoing the epithelial to mesenchymal transition (EMT), a process associated with the metastasis of tumor cells. During this transition, the markers EpCAM, vimentin, and cytokeratin are presumed to change their expression. The most popular, current technology used to capture CTC is the CellSearch® method. This immunoaffinity method relies on the expression of EpCAM, an epithelial cell marker, to capture CTC and may miss cells due to varied cell phenotypes and long processing times. The device used to capture the cells in this study was size based and therefore could capture cells without a positive EpCAM expression. NCI-H441 (lung cancer) cells were spiked into blood for various amounts of time before filtration, ranging from 0 to 24 hours. Cells were filtered, fixed, and stained using immunofluorescent antibodies. The intensity of the staining was related to expression of the marker. There was a significant decrease in EpCAM, cytokeratin, and vimentin expression from 0 to 4 hours, but expression remained similar from 4 to 24 hours. This illustrates that the phenotype of the cell is potentially changing due to either the EMT, the chemistry of the blood, or the life of the cell. The results suggest that immunoaffinity based capture methods may not provide correct information about the patient’s cancer due to the time between blood draw and CTC capture and the cells’ altered phenotypes. Future research should be performed to provide additional time points to determine the approximate time before this alteration, and therefore a desired maximum processing time.