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Ewing’sarcoma, the second most frequent bone tumor of young adults, is characterized by the specific translocation t(11;22), leads to the formation of the aberrant transcription factor EWS-FLI1. From the histological point of view, Ewing cells are undifferentiated and their cellular origin, a knowledge required for the complete understanding of this sarcoma, is currently unknown.

In order to identify the cellular origin of Ewing tumors, we made the hypothesis that EWS-FLI1-inihbited Ewing cells should re-express some markers specific of their original tissue and thus recover, at least in part, their primary phenotype. In a first step, we compared expression profiles of Ewing cell lines inhibited for EWS-FLI1 with those of a collection of normal tissues, including mesenchymal stem cells. Principal component analysis of these different profiles unambiguously showed that EWS-FLI1-inhibited cells cluster together with mesenchymal tissues. In a second step, aware of the close relation between the expression profiles of EWS-FLI1-inhibited cells and mesenchymal stem cells, we performed mesenchymal differentiation experiments on Ewing cells. For this purpose, we constructed a stable cell line expressing a shRNA directed against EWS-FLI1 that is inducible upon treatment with doxycycline. Using this original system, we could show that EWS-FLI1-inhibited cells expressed specific markers from either adipocytes, osteocytes or chondrocytes lineages when grown in the corresponding differentiation medium. Moreover, the positive staining of the cells either by Oil-Red-O (detection of lipidic vesicles) or by the Von-Kossa protocol (detection of a calcified matrix) demonstrates a commitment of Ewing cells inhibited for EWS-FLI1 towards the adipocytes and osteocytes lineages, respectively. Finally, we could show that these cells could also be phenotypically characterized as mesenchymal stem cells by their cell surface antigens expression profile.

Altogether, these results show that Ewing cells inhibited for EWS-FLI1 are able to recover some of the differentiation features characteristic of mesenchymal stem cells, supporting the hypothesis of a mesenchymal origin of Ewing tumors.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA