Absence of CD71 Transferrin Receptor Characterizes Human Gastric Adenosquamous Carcinoma Stem Cells
Department of Gastroenterological Surgery, Graduate School of Medicine, Osaka University, Osaka, Japan. Annals of Surgical Oncology
(Impact Factor: 3.93).
04/2011; 19(4):1357-64. DOI: 10.1245/s10434-011-1739-7
Although the importance of cancer stem cells (CSCs) in overcoming resistance to therapy and metastasis has recently been reported, the role of CSCs in gastric cancer remains to be elucidated.
MKN-1 cells were used to study markers of CSCs in gastric adenosquamous carcinoma, as these cells are suitable for determining multidifferentiation ability. Changes in expression of CD44, CD49f, CD133, and CD71 following 5-fluorouracil (5-FU) treatment were assessed.
After 5-FU treatment, only the CD71- fraction was significantly increased. Investigation of CD71 indicated that the CD71- cell fraction was present in the G1/G0 cell cycle phase and showed high resistance to the anticancer agent 5-FU. Limiting dilution and serial transplantation assays revealed the CD71- cell fraction to have higher tumorigenicity than the CD71+ cell fraction. The CD71- cell fraction showed multipotency to adenocarcinoma and squamous cell carcinoma. A three-dimensional (3D) invasion assay and immunohistochemical analysis showed CD71- cells to be highly invasive and to exist in the invasive fronts of cancer foci.
The present study suggests that use of CD71- as a marker for adenosquamous carcinoma may provide a useful model for studying CSCs.
Available from: Claire H Seedhouse
- "As Annexin V is a highly sensitive marker for early apoptosis in primary AML cells, we looked for a cell surface dormancy marker that could be used in conjunction with Annexin V to measure apoptosis in dormant compared to proliferating cells. CD71 (the transferrin receptor) is absent in un-stimulated peripheral blood lymphocytes, in some cancer stem cells and in long term culture-initiating cells from normal bone marrow [36,37,47]. Analysis of patient samples co-labelled with CD71 and Ki-67 indicated that CD71 is not expressed in dormant AML blasts (Figure 5). "
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Dormant cells are characterised by low RNA synthesis. In contrast, cancer cells can be addicted to high RNA synthesis, including synthesis of survival molecules. We hypothesised that dormant cancer cells, already low in RNA, might be sensitive to apoptosis induced by RNA Polymerase II (RP2) inhibitors that further reduce RNA synthesis.
We cultured leukaemia cells continuously in vitro in the presence of an mTOR inhibitor to model dormancy. Apoptosis, damage, RNA content and reducing capacity were evaluated. We treated dormancy-enriched cells for 48 hours with the nucleoside analogues ara-C, 5-azacytidine and clofarabine, the topoisomerase targeting agents daunorubicin, etoposide and irinotecan and three multikinase inhibitors with activity against RP2 - flavopiridol, roscovitine and TG02, and we measured growth inhibition and apoptosis. We describe use of the parameter 2 × IC50 to measure residual cell targeting. RNA synthesis was measured with 5-ethynyl uridine. Drug-induced apoptosis was measured flow cytometrically in primary cells from patients with acute myeloid leukaemia using a CD34/CD71/annexinV gating strategy to identify dormant apoptotic cells.
Culture of the KG1a cell line continuously in the presence of an mTOR inhibitor induced features of dormancy including low RNA content, low metabolism and low basal ROS formation in the absence of a DNA damage response or apoptosis. All agents were more effective against the unmanipulated than the dormancy-enriched cells, emphasising the chemoresistant nature of dormant cells. However, the percentage of cell reduction by RP2 inhibitors at 2 × IC50 was significantly greater than that of other agents. RP2 inhibitors strongly inhibited RNA synthesis compared with other drugs. We also showed that RP2 inhibitors induce apoptosis in proliferating and dormancy-enriched KG1a cells and in the CD71neg CD34pos subset of primary acute myeloid leukaemia cells.
We suggest that RP2 inhibitors may be a useful class of agent for targeting dormant leukaemia cells.
Available from: europepmc.org
- "and CD90 + identifying them. These GCSCs possess a high migratory and invasive potential and enhanced drug resistance capa- bility   . However, the mechanisms involved remain largely unknown. "
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ABSTRACT: Gastric cancer stem-like cells (GCSCs) have been identified to possess the ability of self-renewal and tumor initiation. However, the mechanisms involved remain largely unknown. Here, we isolated and characterized the GCSCs by side population (SP) sorting procedure and cultured sphere cells (SC) from human gastric cancer cell lines SGC-7901, BGC-823, MGC-803, HGC-27 and MKN-28. The sorting and culture assay revealed that SP cells proliferated in an asymmetric division manner. In addition, SP cells exhibited a higher potential of spheroid colony formation and greater drug resistance than non-SP cells (NSP). Moreover, the SC were found with enhanced capabilities of drug resistance in vitro and tumorigenicity in vivo. Sox2 mRNA and protein was highly and significantly overexpressed in the SP cells and SC. Importantly, downregulation of Sox2 with siRNA obviously reduced spheroid colony formation and doxorubicin efflux, as well as increased apoptosis rate in sphere cells in vitro and suppressed tumorigenicity in vivo. These results suggest that both SP cells and cultured SC enrich with GCSCs and that Sox2 plays a pivotal role in sustaining stem cell properties and might be a potential target for gastric cancer therapy.
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ABSTRACT: Gastric cancer (GC) remains one of the most common cancers worldwide. Its prevalence is still on the rise in the developing countries due to the ageing population. The cancer stem cell (CSC) theory provides a new insight into the interpretation of tumor initiation, aggressive growth, recurrence, and metastasis of cancer, as well as the development of new strategies for cancer treatment. This review will focus on the progress of biomarkers and signaling pathways of CSCs, the complex crosstalk networks between the microenvironment and CSCs, and the development of therapeutic approaches against CSCs, predominantly focusing on GC.
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