Stable interference of EWS-FLI1 in an Ewing sarcoma cell line impairs IGF-1/IGF-1R signalling and reveals TOPK as a new target

D Herrero-Martín; D Osuna; J L Ordóñez; V Sevillano; A S Martins; C Mackintosh; M Campos; J Madoz-Gúrpide; A P Otero-Motta; G Caballero; A T Amaral; D H Wai; Y Braun; M Eisenacher; K-L Schaefer; C Poremba; E de Alava

doi:10.1038/sj.bjc.6605104

Stable interference of EWS-FLI1 in an Ewing sarcoma cell line impairs IGF-1/IGF-1R signalling and reveals TOPK as a new target

Br J Cancer. 2009 Jul 7;101(1):80-90. doi: 10.1038/sj.bjc.6605104. Epub 2009 Jun 2.

Authors

D Herrero-Martín¹, D Osuna, J L Ordóñez, V Sevillano, A S Martins, C Mackintosh, M Campos, J Madoz-Gúrpide, A P Otero-Motta, G Caballero, A T Amaral, D H Wai, Y Braun, M Eisenacher, K-L Schaefer, C Poremba, E de Alava

Affiliation

¹ Molecular Pathology Program, Centro de Investigación del Cáncer-IBMCC (USAL-CSIC), Campus Unamuno s/n, Salamanca, Spain. david.herrero@kispi.uzh.ch

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.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Apoptosis / physiology
Cell Line, Tumor
Cell Movement / physiology
Down-Regulation
Female
Humans
Insulin-Like Growth Factor I / antagonists & inhibitors
Insulin-Like Growth Factor I / biosynthesis
Insulin-Like Growth Factor I / genetics
Insulin-Like Growth Factor I / metabolism*
Mice
Mice, Inbred NOD
Mice, SCID
Mitogen-Activated Protein Kinase Kinases
Oncogene Proteins, Fusion / antagonists & inhibitors*
Oncogene Proteins, Fusion / genetics
Oncogene Proteins, Fusion / metabolism
Protein Serine-Threonine Kinases / biosynthesis*
Protein Serine-Threonine Kinases / genetics
Protein Serine-Threonine Kinases / metabolism
Proto-Oncogene Protein c-fli-1
RNA Interference
RNA-Binding Protein EWS
Receptor, IGF Type 1 / antagonists & inhibitors
Receptor, IGF Type 1 / biosynthesis
Receptor, IGF Type 1 / genetics
Receptor, IGF Type 1 / metabolism*
Sarcoma, Ewing / enzymology
Sarcoma, Ewing / genetics
Sarcoma, Ewing / metabolism*
Sarcoma, Ewing / pathology
Signal Transduction
Transcription Factors / antagonists & inhibitors*
Transcription Factors / genetics
Transcription Factors / metabolism

Substances

EWS-FLI fusion protein
Oncogene Proteins, Fusion
Proto-Oncogene Protein c-fli-1
RNA-Binding Protein EWS
Transcription Factors
Insulin-Like Growth Factor I
Receptor, IGF Type 1
Protein Serine-Threonine Kinases
Mitogen-Activated Protein Kinase Kinases
PDZ-binding kinase