Key result
Both EWS/WT1-KTS and EWS/WT1 + KTS isoforms demonstrated oncogenic potential, enhancing cell proliferation and survival in murine embryonic fibroblasts only when p53 function was lost.
Why the study?
The oncogenic properties of the two splice variants of EWS/WT1 in desmoplastic small round cell tumor and their interaction with p53 status were not fully understood.
Both isoforms of EWS/WT1 have oncogenic potential in MEFs with loss of p53, providing a novel link between EWS/WT1 and Wnt-pathway signaling in DSRCT.
p53 loss unmasks EWS/WT1 oncogenic activity in MEFs; leaves open p53-Wnt interactions as therapeutic targets in DSRCT.
BACKGROUND: Desmoplastic small round cell tumor (DSRCT) is characterized by the presence of a fusion protein EWS/WT1, arising from the t (11;22) (p13;q12) translocation. Here we examine the oncogenic properties of two splice variants of EWS/WT1, EWS/WT1-KTS and EWS/WT1 + KTS. METHODS: We over-expressed both EWS/WT1 variants in murine embryonic fibroblasts (MEFs) of wild-type, p53+/- and p53-/- backgrounds and measured effects on cell-proliferation, anchorage-independent growth, clonogenicity after serum withdrawal, and sensitivity to cytotoxic drugs and gamma irradiation in comparison to control cells. We examined gene expression profiles in cells expressing EWS/WT1. Finally we validated our key findings in a small series of DSRCT. RESULTS: Neither isoform of EWS/WT1 was sufficient to transform wild-type MEFs however the oncogenic potential of both was unmasked by p53 loss. Expression of EWS/WT1 in MEFs lacking at least one allele of p53 enhanced cell-proliferation, clonogenic survival and anchorage-independent growth. EWS/WT1 expression in wild-type MEFs conferred resistance to cell-cycle arrest after irradiation and daunorubicin induced apoptosis. We show DSRCT commonly have nuclear localization of p53, and copy-number amplification of MDM2/MDMX. Expression of either isoform of EWS/WT1 induced characteristic mRNA expression profiles. Gene-set enrichment analysis demonstrated enrichment of WNT pathway signatures in MEFs expressing EWS/WT1 + KTS. Wnt-activation was validated in cell lines with over-expression of EWS/WT1 and in DSRCT. CONCLUSION: In conclusion, we show both isoforms of EWS/WT1 have oncogenic potential in MEFs with loss of p53. In addition we provide the first link between EWS/WT1 and Wnt-pathway signaling. These data provide novel insights into the function of the EWS/WT1 fusion protein which characterize DSRCT.
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Bandopadhayay et al. (2013) studied Desmoplastic small round cell tumor (n=15). EWS/WT1-KTS and EWS/WT1 + KTS over-expression vs. eGFP control was evaluated on Oncogenic transformation (cell proliferation, clonogenic survival, anchorage-independent growth). Both EWS/WT1-KTS and EWS/WT1 + KTS isoforms demonstrated oncogenic potential, enhancing cell proliferation and survival in murine embryonic fibroblasts only when p53 function was lost.
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