Circulating tumor cells exhibit stem cell characteristics in an orthotopic mouse model of colorectal cancer.

Sebastian Schölch,Lahiri K Nanduri,Nuh N Rahbari,Ulrich Bork,Christoph Kahlert,Markus W Büchler,Thomas Niemietz,Alexander M Betzler,Naoki Iwata,Christoph Reissfelder,Anka Swiersy,May-Linn Thepkaysone,Jürgen Weitz,Sebastián A García

doi:10.18632/oncotarget.8373

Abstract

The prognosis of colorectal cancer (CRC) is closely linked to the occurrence of distant metastases, which putatively develop from circulating tumor cells (CTCs) shed into circulation by the tumor. As far more CTCs are shed than eventually metastases develop, only a small subfraction of CTCs harbor full tumorigenic potential. The aim of this study was to further characterize CRC-derived CTCs to eventually identify the clinically relevant subfraction of CTCs.We established an orthotopic mouse model of CRC which reliably develops metastases and CTCs. We were able to culture the resulting CTCs in vitro, and demonstrated their tumor-forming capacity when re-injected into mice. The CTCs were then subjected to qPCR expression profiling, revealing downregulation of epithelial and proliferation markers. Genes associated with cell-cell adhesion (claudin-7, CD166) were significantly downregulated, indicating a more metastatic phenotype of CTCs compared to bulk tumor cells derived from hepatic metastases. The stem cell markers DLG7 and BMI1 were significantly upregulated in CTC, indicating a stem cell-like phenotype and increased capacity of tumor formation and self-renewal. In concert with their in vitro and in vivo tumorigenicity, these findings indicate stem cell properties of mouse-derived CTCs.In conclusion, we developed an orthotopic mouse model of CRC recapitulating the process of CRC dissemination. CTCs derived from this model exhibit stem-cell like characteristics and are able to form colonies in vitro and tumors in vivo. Our results provide new insight into the biology of CRC-derived CTCs and may provide new therapeutic targets in the metastatic cascade of CRC.

Highlights

Distant metastases rather than the primary tumor cause tumor-related death in over 90% of colorectal cancer (CRC) patients [1]
As tumor cell dissemination during the implantation could obscure the results of the subsequent circulating tumor cells (CTCs) analysis, we developed a method by which the injection of tumor cells into the cecal wall is highly controlled and reproducible as it is performed by a precision micro pump and visually controlled through a microscope
HCT116 grows at a significantly higher speed in vitro; we chose to inject a lower number of HCT116 tumor cells to allow the tumor to form and seed distant metastases before peritoneal dissemination or bowel obstruction occur

Summary

Introduction

Distant metastases rather than the primary tumor cause tumor-related death in over 90% of colorectal cancer (CRC) patients [1]. The biologic origin of metastasis are circulating tumor cells (CTCs), which detach from the tumor, survive in circulation, attach to the endothelium within the target organ, invade the parenchyma and form new tumors [2]. As most distant metastases strikingly resemble the primary tumor both genetically and phenotypically, the changes a tumor cell must undergo to be able to spread to a distant organ must be transient and reversible [14,15,16]. Knowledge of these phenotypic alterations of CTCs may potentially provide new targets to delay or even prevent the formation of new metastases

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