Inhibition of cyclin-dependent kinase 1-induced cell death in neuroblastoma cells through the microRNA-34a-MYCN-survivin pathway.
BACKGROUND
Neuroblastoma is the most common pediatric solid extracranial malignancy. Increased expression of MYCN, a member of Myc family, is associated with a poor prognosis in patients with neuroblastoma. Cyclin-dependent kinase 1 (CDK1) is known to be critical to the survival of Myc-overexpressing neoplasms. In this study, we investigated the effects of CDK1 inhibition on the viability of neuroblastoma cells and the underlying molecular mechanisms.


METHODS
We assessed the effects of CDK1 inhibitors on the viability of neuroblastoma cells and explored MYCN-associated mechanisms by using real-time reverse-transcription polymerase chain reaction, chromatin immunoprecipitation, and micro-RNA quantification.


RESULTS
CDK1 inhibitors induced cytotoxicity and apoptosis and decreased expression of MYCN and survivin in neuroblastoma cells. Knockdown of MYCN mRNA expression caused significant cell death and transcomplementation of MYCN increased cell viability in neuroblastoma cells treated with CDK1 inhibitors. Furthermore, MYCN mRNA was inhibited by CDK1 inhibitor at the posttranscriptional level. The expression of survivin was also responsive to the change of MYCN levels, and according to the chromatin immunoprecipitation assay, MYCN bound the promoter region of the survivin gene. CDK1 inhibition caused elevation of microRNA-34a (miR-34a), and anti-miR-34a not only restored the expression of MYCN but also increased cell survival of neuroblastoma cells treated with CDK1 inhibitor.


CONCLUSIONS
CDK1 inhibition-induced cell death was dependent, although not completely, on the decreased level of MYCN- and miR-34a-mediated, CDK1 inhibition-induced down-regulation of MYCN, which thereby decreased the transcriptional activation of MYCN on the survivin promoter. We propose that CDK1 inhibition-induced cell death of neuroblastoma cells occurs through the miR-34a-MYCN-survivin pathway.
Department of Surgery, Far Eastern Memorial Hospital, Pan-Chiao, New Taipei, Taiwan.
Yun
Y
Chen
Ya-Hui
YH
Tsai
Sheng-Hong
SH
Tseng
D000970:Antineoplastic Agents / Q000494:pharmacology* / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects*; D014408:Biomarkers, Tumor / Q000378:metabolism*; D015153:Blotting, Western; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D047369:Chromatin Immunoprecipitation; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D005434:Flow Cytometry; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000378:metabolism; D035683:MicroRNAs / Q000378:metabolism; D000071447:N-Myc Proto-Oncogene Protein; D009447:Neuroblastoma / Q000188:drug therapy* / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D015513:Oncogene Proteins / Q000378:metabolism; D011687:Purines / Q000494:pharmacology* / Q000627:therapeutic use; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077546:Roscovitine; D000077022:Survivin
C471843:6-((3-chloro)anilino)-2-(isopropyl-2-hydroxyethylamino)-9-isopropylpurine; D000970:Antineoplastic Agents; C107489:BIRC5 protein, human; D014408:Biomarkers, Tumor; D051034:Inhibitor of Apoptosis Proteins; C522425:MIRN34 microRNA, human; C459801:MYCN protein, human; D035683:MicroRNAs; D000071447:N-Myc Proto-Oncogene Protein; D009687:Nuclear Proteins; D015513:Oncogene Proteins; D011687:Purines; D000077022:Survivin; D000077546:Roscovitine; D016203:CDC2 Protein Kinase