Abstract LB-358: A mechanistic study on the metastasis inducing function of FUS-CHOP fusion protein in liposarcoma
Authors: Nitin Patil, Kabeer Suhail Ahmed Rasheed, Jörg Hendrik Leupold, Matthias Schwarzbach, Heike Allgayer
Journal: Cancer Research (2011)

Abstract Myxoid liposarcoma exhibits a stable reciprocal translocation, leading to expression of FUS-CHOP chimeric oncogenic protein. Furthermore, studies have shown that this event promotes tumor formation in nude mice. However, the molecular mechanisms associated with FUS-CHOP as a putative transcription factor, and its ability to promote invasion and metastasis have never been investigated in detail. The present project was conducted to determine the role of FUS-CHOP in mediating invasion and metastasis, and to define potential target genes and molecular mediators. We found that stable overexpression of FUS-CHOP-I protein in SW872 liposarcoma and HT1080 fibrosarcoma cells leads to increased migration and invasion in matrigel assays. These results were supported by cell proliferation assays, showing no significant difference between FUS-CHOP stably over-expressing cells compared with mock-transfected cells. Based on our finding from microarray data gained from 3 resected liposarcoma patients, we investigated the expression of MMPs and observed that MMP-2, 9, and 7 mRNA were significantly up regulated (&amp;gt;3 fold), in both FUS-CHOP stably overexpressing cell lines. Comparing FUS-CHOP positive (n=14) and negative (n=17) liposarcoma patients, we confirmed MMP-2 expression to be significantly higher (p=0.002) in FUS-CHOP positive samples. Luciferase experiments using the basal promoter regions of MMP-2, −7, and −9 demonstrated that an overexpression of FUS-CHOP enhances the promoter activity of all 3 genes, with MMP-2 showing the highest inducibility. Similarly a 4X-AP1 Luc-reporter construct in both cell lines showed a more than 2-fold induction of reporter activity, indicating that FUS-CHOP is able to induce AP1-related transactivation. In contrast, a 6X-NFkB construct showed decreased reporter activity. In addition, chromatin-immunoprecipitation (ChIP) assays revealed an in vivo binding of FUS-CHOP to AP1 and C-EBP-β sites within the MMP-2 promoter after FUS-CHOP transfection. Finally, in vivo chicken embryo metastasis assays (CAM) showed a significantly increased metastasis to lungs and liver of the embryos in FUS-CHOP overexpressing liposarcoma cell lines. Furthermore, primary tumours excised from the upper CAM of this model showed an increase in MMP-2,−7 and −9 expression for both cell lines used. No significant increase in tumor weight was measured, suggesting that the ability of FUS-CHOP to induce metastasis is proliferation-independent. Taken together, these results for the first time suggest that FUS-CHOP overexpression leads to increased migration, invasion and metastasis of liposarcoma and fibrosarcoma cells, in part due to the transcriptional regulation of MMPs. Ongoing studies in our laboratory will elucidate more precise molecular mechanisms related to FUS-CHOP as a transcription factor. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-358. doi:10.1158/1538-7445.AM2011-LB-358