ABSTRACT
Acquired resistance to classical chemotherapeutics is a major obstacle in cancer treatment. Doxorubicin is frequently used in breast cancer therapy either as single-agent or in combination with other drugs like docetaxel and cyclophosphamide. All these chemotherapies have in common that they are administered sequentially and often result in chemoresistance. Here, we mimicked this pulse therapy of breast cancer patients in an in vitro cell culture model, where the epithelial breast cancer cell line BT474 was sequentially treated with doxorubicin for several treatment cycles. In consequence, we obtained chemoresistant cells displaying a mesenchymal-like phenotype with decreased levels of miR-200c. To investigate the involvement of miR-200c in resistance formation, we inhibited and overexpressed miR-200c in different cell lines. Thereby, the cells were rendered more resistant or susceptible to doxorubicin treatment. Moreover, the receptor tyrosine kinase TrkB and the transcriptional repressor Bmi1 were identified as miR-200c targets mediating the drug resistance. Hence, we provide a mechanism of acquired resistance to doxorubicin that is caused by the loss of miR-200c. Along with this, our study demonstrates the complex network of microRNA mediated chemoresistance highlighting the challenges in cancer therapy and the importance of novel microRNA-modulating anticancer agents.
Subject(s)
Antineoplastic Agents/pharmacology , Breast Neoplasms/metabolism , Doxorubicin/pharmacology , MicroRNAs/metabolism , Polycomb Repressive Complex 1/metabolism , Receptor, trkB/metabolism , Breast Neoplasms/genetics , Cell Line, Tumor , Evolution, Molecular , Gene Expression Regulation, Neoplastic , Humans , Immunoblotting , MicroRNAs/genetics , Polycomb Repressive Complex 1/genetics , Receptor, trkB/genetics , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
A major obstacle in the successful treatment of cancer is the occurrence of chemoresistance. Cancer cells surviving chemotherapy and giving rise to a recurrence of the tumor are termed cancer stem cells and can be identified by elevated levels of certain stem cell markers. Eradication of this cell population is a priority objective in cancer therapy. Here, we report elevated levels of stem cell markers in MCF-7 mammospheres. Likewise, an upregulation of HER2 and its differential expression within individual cells of mammospheres was observed. Sorting for HER2(high) and HER2(low) cells revealed an upregulation of stem cell markers NANOG, OCT4 and SOX2 in the HER2(low) cell fraction. Accordingly, HER2(low) cells also showed reduced proliferation, ductal-like outgrowths and an increased number of colonies in matrigel. Xenografts from subcutaneously injected HER2(low) sorted cells exihibited earlier onset but slower growth of tumors and an increase in stem cell markers compared to tumors developed from the HER2(high) fraction. Treatment of mammospheres with salinomycin reduced the expression of SOX2 indicating a selective targeting of cancer stem cells. Trastuzumab however, did not reduce the expression of SOX2 in mammospheres. Furthermore, a combinatorial treatment of mammospheres with trastuzumab and salinomycin was superior to single treatment with each drug. Thus, targeting HER2 expressing tumors with anti-HER2 therapies will not necessarily eliminate cancer stem cells and may lead to a more aggressive cancer cell phenotype. Our study demonstrates efficient killing of both HER2 positive cells and cancer stem cells, hence opening a possibility for a new combinatorial treatment strategy.
