Putative Mesothelioma Stem Cells May be the Key Target to Inhibit Cancer Cell Repopulation during the Intervals of Chemoradiation in Murine Models

Cancer cell repopulation between cycles of chemotherapy is a major factor limiting the efficacy of treatment. Cancer stem cells (CSC) may play critical roles in this process. Our hypothesis is to identify mesothelioma stem cells (MSC) and then make it feasible to design specific target therapy against MSC and improve the efficacy of malignant mesothelioma.

Methods:Murine mesothelioma AB12 and RN5 cells with or without chemoradiation were used to compare tumor incidence in vivo and evaluate gene expression in vitro by real time PCR. Then we developed RN5-EOS-Puro-R cell line by transducing the EOS (Early transposon promoter and Oct-4 (Pou5f1) and Sox2 enhancers) into RN5 cells with lentiviral vector. RN5-EOS knock-in cells with GFP reporter can be selected by puromycin so as to achieve a highly enriched population of putative MSC. Characterization of MSC and RN5-EOS parental cells were investigated. Gene profile was evaluated by microarray assay to screen potential MSC markers.

Results:Tumor growth delay of murine mesothelioma AB12 was achieved after each cycle of cisplatin, however, tumors grew back up rapidly due to cancer cell repopulation between courses of chemotherapy. Strikingly, 10 times less number of irradiated cells in both cell lines can lead to similar tumor incidence and growth rate than untreated cells. The expression of cancer stem cell-associated genes such as CD24, CD133, CD90 and uPAR were up-regulated dramatically after chemoradiation. Highly enriched MSC after selection with puromycin displayed increasing GFP population and had typical property of stemness. Comparatively, the proportion of MSC significantly increased after RN5-EOS parental cells were treated with either chemotherapy, or γ-ray radiation, or in combination, while MSC showed more resistant to the above treatments. Gene profile identified a group of genes that are most likely MSC specific.

Conclusion:Putative MSC possess property of stemness showing more resistance to chemoradiation, suggesting that MSC may play critical roles in cancer cell repopulation. Further identification of selected genes may be used to design novel target therapy against MSC in murine mesothelioma models. Targeting MSC would be able to eliminate cancer cell repopulation and may be suitable to other types of cancer as well.

Keywords: Mesothelioma stem cell (MSC); Enriched RN5-EOS-Puro-R cells; Cancer cell repopulation; Microarray; Chemoradiation