Abstract A20: Protein kinase C beta II is crucial for Ewing sarcoma cell survival

Abstract Ewing sarcoma, the second most frequent bone tumor of young adults, is most frequently characterized by a specific translocation t(11;22) leading to the formation of the aberrant transcription factor EWS-FLI1. By analyzing transcriptional profiles of Ewing tumors with other pediatric tumors, we identified genes that were highly and very specifically over-expressed in Ewing sarcomas, as compared to the others. Beside already identified EWS-FLI1 targets like NKX2-2 or NR0B1, we focused our attention on a novel potential EWS-FLI1 target gene: the protein kinase C beta (PRKCB). Firstly, we confirmed at the protein level, that PRKCB is highly expressed in different Ewing cell lines. Secondly, EWS-FLI1 shRNA inhibition experiments in different Ewing cell lines demonstrated that the PRKCB over-expression is dependent on the expression of the fusion protein. Thirdly, using luciferase reporter experiments together with chromatin immunoprecipitation, we could show that EWS-FLI1 can bind directly to the PRKCB promoter and regulate its activity, demonstrating that PRKCB is a new direct target of EWS-FLI1. Finally, we investigated the role of the two different PRKCB isoforms in Ewing cells by inhibiting either both or the second one only by knock-down experiments. Both strategies led to a dramatic decrease of cell proliferation together with a massive cell death, indicating that at least PRKCB isoform 2, which seems to be the most expressed isoform in Ewing cells, is crucial for Ewing cell survival. We obtained the same results while using specific inhibitor compounds of PRKCB. Altogether, these results show that protein kinase C beta is highly expressed in Ewing tumors and that its inhibition could be of major interest for developing new therapeutic strategies. Citation Information: Cancer Res 2009;69(23 Suppl):A20.