1. ¹Molecular Pathology Program, Centro de Investigación del Cáncer-IBMCC (USAL-CSIC), Campus Unamuno s/n, Salamanca 37007, Spain
2. ²Department of Pathology, Children's Hospital Los Angeles, 4650 Sunset Boulevard, Los Angeles, CA 90027, USA
3. ³Institute of Pathology, Heinrich-Heine-University, Moorenstrasse 5, Duesseldorf 40225, Germany
4. ⁴Department of Integrated Functional Genomics, University of Muenster, Von-Esmarch Strasse 56, Muenster 48149, Germany

Correspondence: Dr D Herrero-Martín, Department of Oncology, University Childrens Hospital, Steinwiesstrasse 75, Zürich 8032, Switzerland. E-mail: david.herrero@kispi.uzh.ch

⁵Current address: Department of Oncology, University Childrens Hospital, Steinwiesstrasse 75, Zürich 8032, Switzerland

⁶Current address: Department of Immunology, Fundación Jiménez Díaz, Av. Reyes Católicos 2, Madrid 28040, Spain

⁷Current address: Medicine Proteome Center, Ruhr University of Bochum, ND04/Nord, Bochum 44780, Germany

⁸Current address: Institute of Pathology Trier, Wissenschaftspark Trier, Max-Planck-Strasse 18+20, Trier 54296, Germany

Received 22 December 2008; Revised 21 April 2009; Accepted 27 April 2009; Published online 2 June 2009.

Abstract

BACKGROUND: Ewing sarcoma is a paradigm of solid tumour -bearing chromosomal translocations resulting in fusion proteins that act as deregulated transcription factors. Ewing sarcoma translocations fuse the EWS gene with an ETS transcription factor, mainly FLI1. Most of the EWS–FLI1 target genes still remain unknown and many have been identified in heterologous model systems.

METHODS: We have developed a stable RNA interference model knocking down EWS–FLI1 in the Ewing sarcoma cell line TC71. Gene expression analyses were performed to study the effect of RNA interference on the genetic signature of EWS–FLI1 and to identify genes that could contribute to tumourigenesis.

RESULTS: EWS–FLI1 inhibition induced apoptosis, reduced cell migratory and tumourigenic capacities, and caused reduction in tumour growth. IGF-1 was downregulated and the IGF-1/IGF-1R signalling pathway was impaired. PBK/TOPK (T-LAK cell-originated protein kinase) expression was decreased because of EWS–FLI1 inhibition. We showed that TOPK is a new target gene of EWS–FLI1. TOPK inhibition prompted a decrease in the proliferation rate and a dramatic change in the cell's ability to grow in coalescence.

CONCLUSION: This is the first report of TOPK activity in Ewing sarcoma and suggests a significant role of this MAPKK-like protein kinase in the Ewing sarcoma biology.