Enrichment and Selective Targeting of Cancer Stem Cells in Colorectal Cancer Cell Lines

Shunqi Wang

doi:10.4172/2161-0436.s2-006

Abstract

Cancer stem cells (CSCs) are the subpopulation of cells within a tumor proposed to be responsible for tumor initiation, relapses, and resistance to chemotherapeutic drugs. Here we optimized sphere culture conditions to isolate and enrich CSCs from colorectal cancer cell line IMCE-Ras. Spheroid cells that developed in culture expressed high levels of putative stem cell markers, and showed stronger anchorage-independent growth abilities and resistance to conventional chemotherapeutic drugs compared with the initial monolayer adherent cells. Xenograft transplantation assays further demonstrated that IMCE-Ras spheroid cells are highly enriched in CSCs. To develop CSC-targeted therapy, we found that the relative percentage of CSC in IMCE-Ras cells was significantly decreased after a short duration exposure to DNA Methylation inhibitor 5-aza-2’-deoxycytidine (5-Aza-dC), indicating that DNA Methylation may be critical for self-renewal and maintenance of CSCs. Indeed, double knockout of DNA methyltransferase 1 (DNMT1) and DNA methyltransferase 3b (DNMT3b) in colon cancer cell line HCT116 resulted in loss of >95% DNA Methylation and complete loss of tumorigenicity both in vitro and in vivo. These data suggest that DNA Methylation is critical for maintenance of the colon CSC population, and a combination of classical chemotherapeutic drugs and DNA Methylation inhibitors may be an effective treatment of colon cancer.

Highlights

Cancer stem cells (CSCs) are defined as “a small subset of cancer cells” within a cancer, which can self-renew and replenish the heterogeneous lineage of cancer cells that comprise the tumor [1]
Yeung et al [15] demonstrated that colorectal cell lines contain CSC populations that can be enriched by the use of an in vitro Matrigelbased differentiation assay together with selection for expression of the CD44 and CD24 cell surface markers
HCT116 cells could form tumorspheres in culture media supplemented with either N2 or F12, tumorsphere forming ability was gradually lost during the serial passages of tumorspheres, indicating that sphere-forming cells can survive for a few passages under these conditions but fail to undergo self-renewal proliferation (Figure 1A)

Summary

Introduction

Cancer stem cells (CSCs) are defined as “a small subset of cancer cells” within a cancer, which can self-renew and replenish the heterogeneous lineage of cancer cells that comprise the tumor [1]. CD133+ HCT116 colon cancer cells were found not to be radio resistant [13] These data challenged the view that CD133 was an effective marker of colon CSCs. In another study, Dalerba et al [14] employed CD44 and epithelial surface antigen (ESA) as stem-cell-specific markers to isolate colorectal CSCs. Recently, Yeung et al [15] demonstrated that colorectal cell lines contain CSC populations that can be enriched by the use of an in vitro Matrigelbased differentiation assay together with selection for expression of the CD44 and CD24 cell surface markers. Despite these lines of evidence demonstrating that CSCs exist in different sources of colon tumor cells, most of these CSC assays are cumbersome and expensive, not ideal for screening and testing of drugs for development of CSC-based therapy

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Results

Conclusion