10IN NEW TOOLS IN THE MOLECULAR DIAGNOSTIC OF LUNG CANCER

Abstract

What can CTCs tell us? Are CTCs a surrogate for tissue? What are the practical applications and current limitations of existing technologies for CTC detection and analysis? What is their clinical significance? Can they be molecularly characterised? How do they compare to cell free DNA detectable in blood? Can ‘stem cells’ be identified in the circulation? Answers to these questions are only just beginning to emerge. CellSearch is a semi-automated system that enriches and counts CTCs by EpCam based immunomagnetic capture. Cells are labelled with DAPI (nuclear stain) and antibodies to cytokeratins and CD45. CTCs are identified as EpCAM, DAPI and cytokeratin positive, and CD45 (white blood cell antigen) negative. ISET (Isolation by Size of Epithelial Tumours) detects CTCs by size exclusion and CD45 immunohistochemistry identifies contaminating white blood cells. Both techniques have limitations for practical application and provide complementary information. Cellsearch is more robust for enumerating CTCs whereas ISET is ‘EpCAM independent’ and may detect EpCAM negative cells that have undergone epithelial-mesenchymal transition. Using Cellsearch, the majority of patients with small cell lung cancer (SCLC) have detectable CTCs compared to less than half of patients with stage IV non-small cell lung cancer (NSCLC). CTCs decline following one cycle of chemotherapy, mirroring radiological response, and demonstrating a pharmacodynamic role. CTC number is an independent prognostic factor for survival. Of greatest interest are molecular characteristics. EpCAM negative CTCs can be detected by ISET, in addition to clusters of CTCs, termed circulating tumour microemboli (CTM). Preliminary studies of expression of markers of epithelial (E-Cadherin, Cytokeratin, EpCAM) and mesenchymal (N-cadherin, Vimentin) transition, apoptosis and proliferation, reveal distinct differences in marker expression for cells comprising CTM compared to solitary CTCs from the same patient. For example, apoptotic morphology is observed in solitary CTCs but not in CTM. Our data suggests that collective cell migration occurs in lung cancer and we hypothesise that this mode of cell travel around the circulation may pose the greatest threat. Disclosure: The author has declared no conflicts of interest. 10IN NEW TOOLS IN THE MOLECULAR DIAGNOSTIC OF LUNG CANCER