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Ewing's Sarcoma

Section analysis with specific colorimetric staining or specific antibody immune-histochemical (IHC) staining of limb elements (consecutive sections) from wildtype and EFPrx1 embryos (E14.5 and E16.5) or postnatal mice at day one (P1) is shown. A) Dimethylmethylene blue (DMMB) staining shows drastic reduction of Glycosaminoglycan, a marker for chondrocytes in mutant embryos. B) An HA-tag specific antibody staining was used to detect the aberrant EF translocation product. EF expression was high at E13.5 and

Ewing sarcoma (ES) is a cancer of bone and soft tissue that is mainly found in pediatric patients. Effective therapy has not been developed due to the lack of relevant test models that faithfully recapitulates ES tumor development. ES is driven by the EWS/FLI1 (EF) translocation product, an aberrant ETS transcription factor. However, it has been challenging to generate a suitable model that harbors the EF translocation. This could suggest that EF is toxic in many cell types and having solely EF translocation expression is insufficient to promote tumor formation. Recent studies have provided evidences that mesenchymal stem cells (MSC) from both mice and humans tolerated EF expression. We introduced a novel ES model that restricts EF expression to early mesenchyme using Prx1Cre (EFPrx1) mice. EFPrx1 newborns display arrested chondrocyte and osteoblast differentiation, causing bone formation defects and polydactyly. However, despite multiple trials transgenic approaches tolerating EF expression lack behind. To generate a functional mouse model which can mimic human ES, we switched to a Cre inducible system. With inducible EF expression in juvenile mice using Prx1CreERT2, we obtained Ewing-like sarcomas that matched human ES gene expression profiles in a significant proportion of induced tumors that were only driven by EF, but loss of Ink4a, a frequent genetic deletion in human ES, accelerated tumor formation.

We found that EWS-FLI1 (EF) expression caused a significant down-regulation of SMAD1/5. Subsequently, expression of SMAD downstream target genes such as DLX5, RUNX2 and OSTERIX were diminished, but the proliferation rate was not significantly changed as analyzed by Ki67 staining (see Figure). 

The RM lab runs a collaborative research effort on Ewing´s Sarcoma, mainly funded by and undertaken together with the LBI-CR Partner Prof. Heinrich Kovar from the CCRI, Vienna. Bone analysis is undertaken in collaboration with Prof. Reinhold Erben, University of Veterinary Medicine, Vienna. 

A New Mouse Model of Metastasizing Ewing Sarcoma: A Representative µCT image of an EF/EF, Prx1CreERT2 mouse showing tumor formation at the bone of the left foreleg. B Percentage of survival of Ewing sarcoma (ES)-bearing mice based on genetic compound mouse models. C Western blot analysis of five representative tumors detects HA tag in murine tumors harboring HA-EWS/FLI1. 

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