Circulating stem cells in colorectal cancer: new tools and potential therapeutic targets.

Aim: Circulating tumour cells (CTCs) are responsible for tumour dissemination. However, low numbers and scarce propensity to expand have hampered the molecular characterization of CTCs in colorectal cancer (CRC) and the identification of cancer stem cells (CSCs) among CTCs. Taking advantage of a new in vivo model of CTC generation, we analysed colorectal CTCs with specific focus on CSCsrelated features. Method: CRC cells derived from surgical specimens are infected with lentiviral particles containing a luciferase-GFP reporter and injected orthotopically into the cecal wall of immunecompromised mice to generate patient-derived xenografts (PDX). Following the appearance of liver metastases, blood is collected via cardiac puncture to isolate CTCs and both metastases and primary tumours are harvested for subsequent analyses. Results: PDX-derived CTCs (PDCs) exhibit stem cell features, being able to generate organoids in vitro and secondary tumours in vivo and contain a subpopulation of cells expressing markers of CSC and/or metastatic cells. Gene expression and proteomic analyses reveal significant differences in pathway activation in PDCs as compared to primary tumour and metastasis-derived cells. Conclusion: Recapitulating the process of CTCs dissemination with primary tumour cells offers an unprecedented opportunity to explore the mechanisms of metastasis formation and to identify new potential therapeutic targets