Ewing’s sarcoma precursors are highly enriched in embryonic osteochondrogenic progenitors
Miwa Tanaka, … , Jun Kanno, Takuro Nakamura
Miwa Tanaka, … , Jun Kanno, Takuro Nakamura
Published June 9, 2014
Citation Information: J Clin Invest. 2014;124(7):3061-3074. https://doi.org/10.1172/JCI72399.
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Research Article Oncology

Ewing’s sarcoma precursors are highly enriched in embryonic osteochondrogenic progenitors

Abstract

Ewing’s sarcoma is a highly malignant bone tumor found in children and adolescents, and the origin of this malignancy is not well understood. Here, we introduced a Ewing’s sarcoma–associated genetic fusion of the genes encoding the RNA-binding protein EWS and the transcription factor ETS (EWS-ETS) into a fraction of cells enriched for osteochondrogenic progenitors derived from the embryonic superficial zone (eSZ) of long bones collected from late gestational murine embryos. EWS-ETS fusions efficiently induced Ewing’s sarcoma–like small round cell sarcoma formation by these cells. Analysis of the eSZ revealed a fraction of a precursor cells that express growth/differentiation factor 5 (Gdf5), the transcription factor Erg, and parathyroid hormone-like hormone (Pthlh), and selection of the Pthlh-positive fraction alone further enhanced EWS-ETS–dependent tumor induction. Genes downstream of the EWS-ETS fusion protein were quite transcriptionally active in eSZ cells, especially in regions in which the chromatin structure of the ETS-responsive locus was open. Inhibition of β-catenin, poly (ADP-ribose) polymerase 1 (PARP1), or enhancer of zeste homolog 2 (EZH2) suppressed cell growth in a murine model of Ewing’s sarcoma, suggesting the utility of the current system as a preclinical model. These results indicate that eSZ cells are highly enriched in precursors to Ewing’s sarcoma and provide clues to the histogenesis of Ewing’s sarcoma in bone.

Authors

Miwa Tanaka, Yukari Yamazaki, Yohei Kanno, Katsuhide Igarashi, Ken-ichi Aisaki, Jun Kanno, Takuro Nakamura

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Figure 5

Differences in gene expression between eSZ and eGP cells.

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Differences in gene expression between eSZ and eGP cells. (A) Comparison...
(A) Comparison of gene expression profiles between eSZ/EWS-FLI1 and eSZ and eSZ/EWS-FLI1 and eGP/EWS-FLI1 48 hours after introduction. Scatter plots of eSZ with (vertical axis) or without EWS-FLI1 (horizontal axis) and eSZ with EWS-FLI1 (vertical axis) or eGP with EWS-FLI1 (horizontal axis) are shown. Red dots indicate probes of present call, and green dots indicate those of absent call. The threshold lines above and below the diagonal indicate y = 2x (2-fold increase) and y = 0.5x (2-fold decrease), respectively. (B) Expression patterns of Dkk2, Prkcb1, and Ezh2 were validated by quantitative RT-PCR. The mean ± SEM of 3 independent experiments are shown. (C) ChIP-PCR for histone modification at Dkk2, Prkcb1, and Ezh2 promoter regions in eSZ, eGP, and murine Ewing’s sarcomas. Rpl30 and Foxa2 were used as controls for active and repressive histone marks, respectively. The mean ± SEM of 3 independent experiments are shown.