1 The presence or absence of tumor cells in the sentinel lymph node (SLN) is the most important prognostic factor in early stage breast cancer. However, the ideal method for SLN examination is still being sought and currently many different protocols are employed.
We have used two methods for tumor cell detection in SLNs; immunomagnetic selection (IMS) using antibodies recognizing the surface proteins EpCAM and Mucin 1, and RT-PCR targeting breast epithelial transcripts (hMAM, AGR2, TFF1 and SBEM). The two methods were used on the same SLN samples, enabling the comparison of results based on the expression of external cell-surface markers (IMS) with those based on intracellular markers (RT-PCR). EpCAM-IMS proved to have highest sensitivity and identified more positive SLNs than the RT-PCR targeting breast epithelial transcripts. Further examination showed that all RT-PCR marker-expressing cells were also positive with EpCAM, but also that EpCAM positive cells lacking expression of epithelial transcripts had increased expression of MMPs, repressors of E-Cadherin, vimentin, EGFR and members of the Wnt signaling pathway (1). Notably, these cells could not be identified using epithelial cytokeratins or mammaglobin as RT-PCR targets, and might represent a subpopulation of mesenchymal like tumor cells with a putative more aggressive phenotype. The results raise important questions regarding existing methods for detection of micrometastatic cells and may have significant clinical implications.
To further evaluate the temporal relationship between the expression of external protein markers (e.g. EpCAM) and internal epithelial/mesenchymal transcriptional markers, we are currently working with model systems using breast epithelial cell lines that are induced to undergo EMT. We aim to establish a timeline for the expression of specific markers used for tumor cell detection, in particular EpCAM but also transcriptional markers.