RT Journal Article
SR Electronic
T1 Cleavage of the Ewing Tumour-specific <em>EWSR1-FLI1</em> mRNA by Hammerhead Ribozymes
JF Anticancer Research
JO Anticancer Res
FD International Institute of Anticancer Research
SP 1901
OP 1908
VO 29
IS 6
A1 REGINA HÜHN
A1 MARTIN S. STAEGE
A1 MANUELA HESSE
A1 BEATE LIEBIG
A1 STEFAN E.G. BURDACH
YR 2009
UL http://ar.iiarjournals.org/content/29/6/1901.abstract
AB Background: Ewing family tumours (EFT) are the second most common bone tumours in children and adolescents. In the majority of EFT, EWSR1-FLI1 (Ewing sarcoma breakpoint region 1-Friend leukaemia virus integration 1) fusion proteins can be detected and EWSR1-FLI1 substantially contributes to the malignant phenotype of EFT. Therefore, inactivation of EWSR1-FLI1 is an interesting strategy for EFT therapy. Materials and Methods: A ribozyme with specificity for EWSR1-FLI1 was developed and the activity in vitro was investigated. Synthetic RNAs corresponding to EWSR1-FLI1 were used as substrates. In addition, the total RNA from EFT cells was used as substrate and the rapid amplification of cDNA ends method for the detection of the cleavage products was used. Results: The ribozyme cleaved the synthetic RNA in a sequence specific manner with high efficiency in vitro. Furthermore, the expected cleavage products were detected after digestion of the total cellular RNA with this ribozyme. A point mutation in the catalytic centre of the ribozyme abolished enzymatic activity. Conclusion: The RNA corresponding to EWSR1-FLI1 is accessible for ribozyme mediated inactivation and ribozymes are able to cleave EWSR1-FLI1 specific RNA in the presence of a high background of normal cellular RNAs.