High impact of miRNA-4521 on FOXM1 expression in medulloblastoma

Daniel Senfter,Robert M Mader,Georg Krupitza,Gerda Ricken,Andreas Peyrl,Irene Slavc,Christine Haberler,Johannes Gojo,Martin Sill,Thomas Czech,Marcel Kool,Mahzeiar Samadaei,Sibylle Madlener

doi:10.1038/s41419-019-1926-1

Abstract

Medulloblastoma, an embryonal tumor of the cerebellum/fourth ventricle, is one of the most frequent malignant brain tumors in children. Although genetic variants are increasingly used in treatment stratification, survival of high-risk patients, characterized by leptomeningeal dissemination, TP53 mutation or MYC amplification, is still poor. FOXM1, a proliferation-specific oncogenic transcription factor, is deregulated in various solid tumors, including medulloblastoma, and triggers cellular proliferation, migration and genomic instability. In tissue samples obtained from medulloblastoma patients, the significant upregulation of FOXM1 was associated with a loss of its putative regulating microRNA, miR-4521. To understand the underlying mechanism, we investigated the effect of miR-4521 on the expression of the transcription factor FOXM1 in medulloblastoma cell lines. Transfection of this microRNA reduced proliferation and invasion of several medulloblastoma cell lines and induced programmed cell death through activation of caspase 3/7. Further, downstream targets of FOXM1 such as PLK1 and cyclin B1 were significantly reduced thus affecting the cell cycle progression in medulloblastoma cell lines. In conclusion, a restoration of miRNA-4521 may selectively suppress the pathophysiological effect of aberrant FOXM1 expression and serve as a targeted approach for medulloblastoma therapy.

Highlights

Brain tumors are the second most frequent neoplasms in childhood following leukemia and represent the leading cause of cancer related deaths in this age group
We focused on miRNA-4521, an uncharted miRNA located on chromosome 17p13.120,21
For Group 4 this difference in expression was statistically significant (Fig. 1b). To corroborate these findings in vitro, we evaluated the expression of miR-4521 in a panel of different MB cell lines belonging to Sonic Hedgehog (SHH) p53 mutated[23] (DAOY and UW228.2) and Group 3 (D341, D425, D458) and Group 3 to 4 (D283) subgroup[5,6,23] and detected a significant downregulation using cerebellum tissue and normal astrocytes as a cellular control (p < 0.001) (Fig. 1c, d)

Summary

Results

Using in-silico analysis of different miRNA databases (targetscan and mirBase), we identified 25 miRNAs located on the p arm of chromosome 17. We included gene array studies using a larger dataset of MB patient tissue samples (Affymetrix 133plus 2.0 Array, n = 423) This dataset confirmed our finding and showed a significant overexpression of FOXM1 compared to normal cerebellum in all MB subgroups (Fig. 2b), but no significant difference between the distinct subgroups (supplementary Fig. 1A). In our patient cohort we observed a significantly higher expression of FOXM1 in patients who developed a tumor recurrence or responded poorly to chemotherapy when compared with patients with a good clinical outcome (supplementary Fig. 1C) To validate this in a larger series, we tested whether FOXM1 expression may correlate with overall survival of the patients using data from the Cavalli dataset[25] (Fig. 2f). Depending on the cell line, upregulation of relative miR-4521 expression

E Chromosome 17

C UCGUGUCC UGAAGGAAUCG

Discussion

Materials and methods