Abstract:
Objective Cancer stem cells (CSCs) are resistant to chemotherapy. Our study aimed to investigate the stem cell-like properties of doxorubicin-resistant osteosarcoma cell line 143B and its correlation with Notch signaling.
Methods We generated doxorubicinresistant osteosarcoma cells by treating them with 2 μm doxorubicin. Stem cell-like properties such as morphology change, Stro-1/CD117 double positive ratio, stem cell-related gene expression, sphere formation efficiency, and EMT character were assessed on day 5 after doxorubicin withdrawal. Notch receptor and its target genes were examined using qPCR and Western blot analysis. The stem cell-like properties of doxorubicin-resistant osteosarcoma cells were assessed when pretreated with Notch inhibitor or vehicle. The anti-tumor effect of Notch inhibitor was tested using a xenograft model.
Results Doxorubicin-resistant osteosarcoma cells were enriched in Stro-1+/CD117+ cells, which showed obvious increased expression of stem cell-related genes, and exhibited enhanced spheroid formation and evident mesenchymal characteristics unlike doxorubicin-sensitive cells. qPCR and Western blot assays showed that Notch intracellular domain 1 (NICD1) and target genes Hes1 and Hey1 were upregulated in doxorubicin-resistant osteosarcoma stem cells compared with those in vehicle cells. Furthermore, pretreatment with a γ-secretase inhibitor (GSI) to prevent Notch signaling enhanced chemo-sensitivity and inhibited doxorubicin-enriched osteosarcoma stem cell activity in vitro. Finally, the Notch inhibitor prevented tumor growth in mice xenograft models.
Conclusion Doxorubicin induced the enrichment of osteosarcoma stem-like cells through Notch signaling, and inactivation of Notch could be useful for overcoming drug resistance and eliminating osteosarcoma.
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