MicroRNA-497 increases apoptosis in MYCN amplified neuroblastoma cells by targeting the key cell cycle regulator WEE1

Hdl Handle:
http://hdl.handle.net/10147/282904
Title:
MicroRNA-497 increases apoptosis in MYCN amplified neuroblastoma cells by targeting the key cell cycle regulator WEE1
Authors:
Creevey, Laura; Ryan, Jacqueline; Harvey, Harry; Bray, Isabella M; Meehan, Maria; Khan, Adnan R; Stallings, Raymond L
Citation:
Molecular Cancer. 2013 Mar 26;12(1):23
Issue Date:
26-Mar-2013
URI:
http://dx.doi.org/10.1186/1476-4598-12-23; http://hdl.handle.net/10147/282904
Abstract:
Abstract Background Neuroblastoma is responsible for 15% of all childhood cancer deaths. Despite advances in treatment and disease management, the overall 5-year survival rates remain poor in high-risk disease (25-40%). MiR-497 was previously identified by our laboratory as a member of a miRNA expression signature, predictive of neuroblastoma patient survival and has been reported as a tumor suppressor in a variety of other cancers. WEE1, a tyrosine kinase regulator of the cell cycle and predicted target of miR-497, has emerged as an oncogene in several cancer types and therefore represents an attractive potential target for novel therapy approaches in high-risk neuroblastoma. Our aim was to investigate the potential tumor suppressive role of miR-497 in high-risk neuroblastoma. Methods Expression levels of miR-497 and WEE1 in tissues and cells were determined using RT-PCR. The effect of miR-497 and siWEE1 on cell viability was evaluated using MTS assays, apoptosis levels were determined using FACS analysis of Annexin V/PI stained cells, and target protein expression was determined using western blot. Luciferase reporter plasmids were constructed to confirm direct targeting. Results were reported as mean±S.E.M and differences were tested for significance using 2-tailed Students t-test. Results We determined that miR-497 expression was significantly lower in high-risk MYCN amplified (MNA) tumors and that low miR-497 expression was associated with worse EFS and OS in our cohort. Over-expression of miR-497 reduced cell viability and increased apoptosis in MNA cells. We identified WEE1 as a novel target for miR-497 in neuroblastoma. Furthermore, our analysis showed that high WEE1 levels are significantly associated with poor EFS and OS in neuroblastoma and that siRNA knockdown of WEE1 in MNA cell lines results in significant levels of apoptosis, supporting an oncogenic role of WEE1 in neuroblastoma. Cisplatin (CDDP) treatment of both miR-497 over-expressing cells and WEE1 inhibited cells, resulted in a significant increase in apoptosis in MNA cells, describing a synergistic effect and therefore a potential therapeutic for high-risk neuroblastoma. Conclusion Our study’s results are consistent with miR-497 being a candidate tumor suppressor in neuroblastoma, through the direct targeting of WEE1. These findings re-enforce the proposal of WEE1 as a therapeutic target in neuroblastoma.
Item Type:
Article
Language:
en
Keywords:
CHILD HEALTH; CANCER; GENETICS

Full metadata record

DC FieldValue Language
dc.contributor.authorCreevey, Lauraen_GB
dc.contributor.authorRyan, Jacquelineen_GB
dc.contributor.authorHarvey, Harryen_GB
dc.contributor.authorBray, Isabella Men_GB
dc.contributor.authorMeehan, Mariaen_GB
dc.contributor.authorKhan, Adnan Ren_GB
dc.contributor.authorStallings, Raymond Len_GB
dc.date.accessioned2013-04-18T10:10:53Z-
dc.date.available2013-04-18T10:10:53Z-
dc.date.issued2013-03-26-
dc.identifier.citationMolecular Cancer. 2013 Mar 26;12(1):23en_GB
dc.identifier.urihttp://dx.doi.org/10.1186/1476-4598-12-23-
dc.identifier.urihttp://hdl.handle.net/10147/282904-
dc.description.abstractAbstract Background Neuroblastoma is responsible for 15% of all childhood cancer deaths. Despite advances in treatment and disease management, the overall 5-year survival rates remain poor in high-risk disease (25-40%). MiR-497 was previously identified by our laboratory as a member of a miRNA expression signature, predictive of neuroblastoma patient survival and has been reported as a tumor suppressor in a variety of other cancers. WEE1, a tyrosine kinase regulator of the cell cycle and predicted target of miR-497, has emerged as an oncogene in several cancer types and therefore represents an attractive potential target for novel therapy approaches in high-risk neuroblastoma. Our aim was to investigate the potential tumor suppressive role of miR-497 in high-risk neuroblastoma. Methods Expression levels of miR-497 and WEE1 in tissues and cells were determined using RT-PCR. The effect of miR-497 and siWEE1 on cell viability was evaluated using MTS assays, apoptosis levels were determined using FACS analysis of Annexin V/PI stained cells, and target protein expression was determined using western blot. Luciferase reporter plasmids were constructed to confirm direct targeting. Results were reported as mean±S.E.M and differences were tested for significance using 2-tailed Students t-test. Results We determined that miR-497 expression was significantly lower in high-risk MYCN amplified (MNA) tumors and that low miR-497 expression was associated with worse EFS and OS in our cohort. Over-expression of miR-497 reduced cell viability and increased apoptosis in MNA cells. We identified WEE1 as a novel target for miR-497 in neuroblastoma. Furthermore, our analysis showed that high WEE1 levels are significantly associated with poor EFS and OS in neuroblastoma and that siRNA knockdown of WEE1 in MNA cell lines results in significant levels of apoptosis, supporting an oncogenic role of WEE1 in neuroblastoma. Cisplatin (CDDP) treatment of both miR-497 over-expressing cells and WEE1 inhibited cells, resulted in a significant increase in apoptosis in MNA cells, describing a synergistic effect and therefore a potential therapeutic for high-risk neuroblastoma. Conclusion Our study’s results are consistent with miR-497 being a candidate tumor suppressor in neuroblastoma, through the direct targeting of WEE1. These findings re-enforce the proposal of WEE1 as a therapeutic target in neuroblastoma.-
dc.language.isoenen
dc.subjectCHILD HEALTHen_GB
dc.subjectCANCERen_GB
dc.subjectGENETICSen_GB
dc.titleMicroRNA-497 increases apoptosis in MYCN amplified neuroblastoma cells by targeting the key cell cycle regulator WEE1en_GB
dc.typeArticleen
dc.language.rfc3066en-
dc.rights.holderLaura Creevey et al.; licensee BioMed Central Ltd.-
dc.description.statusPeer Reviewed-
dc.date.updated2013-04-12T11:03:46Z-
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