Ependymoma stem cells are highly sensitive to temozolomide in vitro and in orthotopic models

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. The efficacies of the two DNA damaging agents were explored in two ependymoma SC lines in vitro and in vivo models. 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 A mu M) induced accumulation of cells in the G(2)/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 p53 beta/gamma 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 beta III-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. Ependymoma growth might be impaired by temozolomide through preferential depletion of a less differentiated, more tumorigenic, MGMT-negative cell population with stem-like properties.