title: Ependymoma stem cells are highly sensitive to temozolomide in vitro and in orthotopic models.
abstract: BACKGROUND
  Ependymoma management remains challenging because of the inherent chemoresistance of this tumor. To determine whether ependymoma stem cells (SCs) might contribute to therapy resistance, we investigated the sensitivity of ependymoma SCs to temozolomide and etoposide.
  
  
  METHODS
  The efficacies of the two DNA damaging agents were explored in two ependymoma SC lines in vitro and in vivo models.
  
  
  RESULTS
  Ependymoma SC lines were highly sensitive to temozolomide and etoposide in vitro, but only temozolomide impaired tumor-initiation properties. Consistently, temozolomide but not etoposide showed significant antitumoral activity on ependymoma SC-driven subcutaneous and orthotopic xenografts by reducing the mitotic fraction. In vitro temozolomide at the EC50 (10 µM) induced accumulation of cells in the G2/M phase that was unexpectedly accompanied by downregulation of p27 and p21 without modulation of full-length p53 (FLp53). Differentiation-committed ependymoma SCs acquired resistance to temozolomide. Inhibition of proliferation was partly due to apoptosis, that occurred earlier in differentiated cells as compared to neurospheres. The activation of apoptosis correlated with an increase in p53β/γ isoforms without modulation of FLp53 under both serum-free and differentiation-promoting media. Incubation of cells in both conditions with temozolomide resulted in increased glioneuronal differentiation exhibiting elevated glial fibrillary acidic protein, galactosylceramidase, and βIII-tubulin expression compared to untreated controls. O(6)-methylguanine DNA methyltransferase (MGMT) transcript levels were very low in SCs, and were increased by treatment and, epigenetically, by differentiation through MGMT promoter unmethylation.
  
  
  CONCLUSIONS
  Ependymoma growth might be impaired by temozolomide through preferential depletion of a less differentiated, more tumorigenic, MGMT-negative cell population with stem-like properties.
authors: Department of Pediatric Oncology, Catholic University, Rome, Italy (D.M., T.S., R.R.); Istituto CSS - Mendel Laboratory, Rome, Italy (G.L.); Department of Biotechnology and Biosciences, Building U3, University of Milan Bicocca, Milan, Italy (E.B.); Institute of Pathology, Catholic University, Rome, Italy (V.A.).
authors: Daniela
authors: D
authors: Meco
authors: Department of Pediatric Oncology, Catholic University, Rome, Italy (D.M., T.S., R.R.); Istituto CSS - Mendel Laboratory, Rome, Italy (G.L.); Department of Biotechnology and Biosciences, Building U3, University of Milan Bicocca, Milan, Italy (E.B.); Institute of Pathology, Catholic University, Rome, Italy (V.A.).
authors: Tiziana
authors: T
authors: Servidei
authors: Department of Pediatric Oncology, Catholic University, Rome, Italy (D.M., T.S., R.R.); Istituto CSS - Mendel Laboratory, Rome, Italy (G.L.); Department of Biotechnology and Biosciences, Building U3, University of Milan Bicocca, Milan, Italy (E.B.); Institute of Pathology, Catholic University, Rome, Italy (V.A.).
authors: Giuseppe
authors: G
authors: Lamorte
authors: Department of Pediatric Oncology, Catholic University, Rome, Italy (D.M., T.S., R.R.); Istituto CSS - Mendel Laboratory, Rome, Italy (G.L.); Department of Biotechnology and Biosciences, Building U3, University of Milan Bicocca, Milan, Italy (E.B.); Institute of Pathology, Catholic University, Rome, Italy (V.A.).
authors: Elena
authors: E
authors: Binda
authors: Department of Pediatric Oncology, Catholic University, Rome, Italy (D.M., T.S., R.R.); Istituto CSS - Mendel Laboratory, Rome, Italy (G.L.); Department of Biotechnology and Biosciences, Building U3, University of Milan Bicocca, Milan, Italy (E.B.); Institute of Pathology, Catholic University, Rome, Italy (V.A.).
authors: Vincenzo
authors: V
authors: Arena
authors: Department of Pediatric Oncology, Catholic University, Rome, Italy (D.M., T.S., R.R.); Istituto CSS - Mendel Laboratory, Rome, Italy (G.L.); Department of Biotechnology and Biosciences, Building U3, University of Milan Bicocca, Milan, Italy (E.B.); Institute of Pathology, Catholic University, Rome, Italy (V.A.).
authors: Riccardo
authors: R
authors: Riccardi
mesh_terms: D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000627:therapeutic use*; D015254:DNA Modification Methylases / Q000235:genetics; D003606:Dacarbazine / Q000031:analogs & derivatives* / Q000627:therapeutic use; D004195:Disease Models, Animal; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004806:Ependymoma / Q000188:drug therapy*; D005909:Glioblastoma / Q000188:drug therapy*; D008819:Mice, Nude; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics / Q000378:metabolism*; D013234:Stem Cells / Q000378:metabolism; D000077204:Temozolomide; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
chemical_list: D018906:Antineoplastic Agents, Alkylating; D016159:Tumor Suppressor Protein p53; D003606:Dacarbazine; D015254:DNA Modification Methylases; D019853:O(6)-Methylguanine-DNA Methyltransferase; D000077204:Temozolomide
keywords: MGMT; differentiation; ependymoma stem cells; orthotopic models; temozolomide