How does a sarcoma circumvent fusion oncoprotein-mediated toxicity?

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Frederic G. Barr, MD, PhD,  University of Pennsylvania School of Medicine
Recipient of the: $25,000 ARIAD and Merck Research Award

Alveolar rhabdomyosarcoma (ARMS) is an aggressive soft tissue sarcoma with a 2;13 translocation that generates a PAX3-FKHR fusion oncoprotein. Introduction of PAX3-FKHR into multiple cell types showed that this oncoprotein exerts growth suppression and/or cell death when expressed at levels comparable to those found in ARMS cells. Subclones from a myoblast population expressing MYCN and an inducible form of PAX3-FKHR showed increasing growth suppression in a plating assay as PAX3-FKHR activity was induced. However, when fibroblasts were added for a focus formation assay, numerous transformed foci formed when PAX3-FKHR was induced. Multiple ARMS cell lines have a similar phenotype of higher focus formation than plating efficiency. In ARMS cells, siRNA-mediated suppression of FGFR4, a PAX3-FKHR downstream target, increases plating efficiency without affecting focus formation. We hypothesize that FGFR4 signaling contributes to the mechanism of PAX3-FKHR-induced toxicity and that stromal cells attenuate the toxic effects of high PAX3-FKHR activity and promote PAX3-FKHR-induced oncogenesis.  To investigate which stromal cells are capable of exerting this effect, we will use our cell culture system to study additional fibroblasts, myoblasts, and mesenchymal cell types associated with preferred sites of ARMS metastasis. We will study the role of FGFR4 signaling in PAX3-FKHR-induced toxicity and oncogenesis by extending our initial RNA interference studies. PCR and western blot approaches will then be used to analyze expression of FGFR4 and its ligands and the activity of downstream MAPK and AKT signaling pathways in these cells. Based on these findings, inhibitors will be selected to examine the role of specific components of the FGFR4 signaling pathway. These studies will provide a comprehensive approach to an important problem in sarcoma pathogenesis and lead to therapeutic strategies to undermine mechanisms by which sarcoma cells circumvent fusion and thereby become sensitized to this toxicity.