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Abstract A193: From MT3 modulation to HDACi treatment in ependymomas
Ependymoma, the third most common brain tumor in the paediatric population, presents limited treatment options, especially in the case of local relapse. Aiming to understand tumor progression, we performed a microarray analysis over 27 relapses of ependymomas using their own samples at diagnosis as...
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Published in: | Molecular cancer therapeutics 2009-12, Vol.8 (12_Supplement), p.A193-A193 |
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Main Authors: | , , , , , , , , , |
Format: | Article |
Language: | English |
Online Access: | Get full text |
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Summary: | Ependymoma, the third most common brain tumor in the paediatric population, presents limited treatment options, especially in the case of local relapse. Aiming to understand tumor progression, we performed a microarray analysis over 27 relapses of ependymomas using their own samples at diagnosis as a control. Consistent down regulation of metallothionein 3 was found in 87% of relapses. MT3 is mostly expressed in the brain. This protein inhibits growth in a variety of cell types; a characteristic not shared by the others members of the MT family. To investigate the involvement of epigenetic factors in MT3 silencing, methylation of CpG islands were analyzed by DNA bisulfite treatment followed by MT3 promoter sequencing. There were no hypermethylated regions over the sequence and the methylation pattern observed could not be correlated with gene expression level. Treatment of primary culture ependymoma cells with 5-Aza-2′-Deoxycytidine, a demethylating agent, failed to induce MT3 transcriptional reactivation, confirming the CpG islands results. On the other hand, a treatment with 300 nMTrichostatin A (TSA), a deacethylation agent, increased MT3 expression by 100 times. Deregulation in epigenetic control by histone modification is known being important in the development of some neoplasias suggesting HDAC inhibitors a new class of promising chemotherapeutic agents, that are currently evaluated in early clinical trials. MTS tests were performed using the HDAC inhibitor SAHA (vorinostat) and 5-Aza-2′-Deoxycytidine. NEM65 ependymoma primary cell culture, ependymoma cell lines Res196, Res253 and Res254; SF188 glioma cell line and DAOY meduloblastoma cells were treated for 72 hours with different concentrations of these agents in three independent experiments. The cell viability of Res196, Res253, Res254, SF188 and DAOY was significantly decreased by 3µM SAHA (30%, 38%, 41%, 20% and 38%, respectively), but NEM65 seemed to be resistant, about 80% of cell viability at 6µM. In contrast, treatment of these cells with 5-Aza-2′-Deoxycytidine proved no effect, neither in cell viability decrease nor in MT3 expression. Trying to understand the differences between sensible cell lines and the resistant NEM65 we performed gene expression analysis by Q-PCR after SAHA treatment. We analysed MT3, Bcl2, Bax and Caspase3 genes. Interestingly, SAHA increased MT3 expression level at least 100 times in the sensible cells, but only 3 times in the resistant NEM65. For the genes involved |
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ISSN: | 1535-7163 1538-8514 |
DOI: | 10.1158/1535-7163.TARG-09-A193 |