LGR5‐Positive Colon Cancer Stem Cells Interconvert with Drug‐Resistant LGR5‐Negative cells and are Capable of Tumor Reconstitution

Abstract

The cancer stem cell (CSC) concept has been proposed as an attractive theory to explain cancer development, and CSCs themselves have been considered as targets for the development of diagnostics and therapeutics. However, many unanswered questions concerning the existence of slow cycling/quiescent, drug-resistant CSCs remain. Here we report the establishment of colon cancer CSC lines, interconversion of the CSCs between a proliferating and a drug-resistant state, and reconstitution of tumor hierarchy from the CSCs. Stable cell lines having CSC properties were established from human colon cancer after serial passages in NOD/Shi-scid, IL-2Rc null (NOG) mice and subsequent adherent cell culture of these tumors. By generating specific antibodies against LGR5, we demonstrated that these cells expressed LGR5 and underwent self-renewal using symmetrical divisions. Upon exposure to irinotecan, the LGR5 1 cells transitioned into an LGR5 2 drug-resistant state. The LGR5 2 cells converted to an LGR5 1 state in the absence of the drug. DNA microarray analysis and immunohistochemistry demonstrated that HLA-DMA was specifically expressed in drug-resistant LGR5 2 cells, and epiregulin was expressed in both LGR5 1 and drug-resistant LGR5 2 cells. Both cells sustained tumor initiating activity in NOG mice, giving rise to a tumor tissue hierarchy. In addition, anti-epiregulin antibody was found to be efficacious in a metastatic model. Both LGR5 1 and LGR5 2 cells were detected in the tumor tissues of colon cancer patients. The results provide new biological insights into drug resistance of CSCs and new therapeutic options for cancer treatment.

LGR5-Targeted Antibody–Drug Conjugate Eradicates Gastrointestinal Tumors and Prevents Recurrence

“…However, as 2 mice showed no recurrent tumors subsequent to ADC treatment, it seems more feasible that the latter is true. In fact, it has been reported that both LGR5-positive intestinal crypt stem cells and colon CSCs can interconvert with LGR5-negative cells (19,41). The conversion of LGR5-positive CSCs was dependent upon exposure to chemotherapy resulting in an LGR5-negative drug-resistant phenotype, although both cell types retained tumor-initiating activity (19).…”

“…In fact, it has been reported that both LGR5-positive intestinal crypt stem cells and colon CSCs can interconvert with LGR5-negative cells (19,41). The conversion of LGR5-positive CSCs was dependent upon exposure to chemotherapy resulting in an LGR5-negative drug-resistant phenotype, although both cell types retained tumor-initiating activity (19). We also observed that anti-LGR5 mAb-treated tumors grew more slowly compared with vehicle and two of them expressed low levels of LGR5.…”

“…A series of studies using mouse models with loss of the tumor suppressor APC (adenomatous polyposis coli) gene have established that LGR5-positive stem cells are the cells-of-origin that drive intestinal and gastric tumor development (6,15). LGR5-high cancer cells have been shown to exhibit the properties of tumor-initiating cells or cancer stem cells (CSC), which retain the capacity for self-renewal (2,(15)(16)(17)(18)(19)(20). CSCs can fuel tumor growth, confer resistance to standard chemotherapy and radiation, and are likely to contribute to recurrent disease (21).…”

Stem Cell Differentiation and Lumen Formation in Colorectal Cancer Cell Lines and Primary Tumors

“…While markers such as LGR5 (33), ephrin type-B receptor 2 (EphB2; ref. 34), and Lrig1 (35) appear to mark well-defined stem cell populations in normal intestine, their expression in tumors does not appear to be entirely specific to cancer stem cells as low clonogenic cells are also marked, at least for EphB2 (31), and LGR5 (36,37). F-actin lumen formation should therefore be useful for defining the cutoff in expression of markers such as LGR5 and EphB2 that define cancer stem cell populations from non-stem cells.…”

KLF5 Regulates the Integrity and Oncogenicity of Intestinal Stem Cells

“…17). We have recently established cancer cell lines from the samples of human colorectal cancers that express Lgr5 and possess CSC properties (18). These Lgr5 þ colonic CSCs, interconvert with drug-resistant Lgr5 À cells after drug treatment and are capable of tumor reconstitution.…”