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MicroRNA-182 promotes leptomeningeal spread of non-sonic hedgehog-medulloblastoma

Abstract

The contribution of microRNAs to the initiation, progression, and metastasis of medulloblastoma (MB) remains poorly understood. Metastatic dissemination at diagnosis is present in about 30% of MB patients, and is associated with a dismal prognosis. Using microRNA expression profiling, we demonstrate that the retinal miR-183–96–182 cluster on chromosome 7q32 is highly overexpressed in non-sonic hedgehog MBs (non-SHH-MBs). Expression of miR-182 and miR-183 is associated with cerebellar midline localization, and miR-182 is significantly overexpressed in metastatic MB as compared to non-metastatic tumors. Overexpression of miR-182 in non-SHH-MB increases and knockdown of miR-182 decreases cell migration in vitro. Xenografts overexpressing miR-182 invaded adjacent normal tissue and spread to the leptomeninges, phenotypically reminiscent of clinically highly aggressive large cell anaplastic MB. Hence, our study provides strong in vitro and in vivo evidence that miR-182 contributes to leptomeningeal metastatic dissemination in non-SHH-MB. We therefore reason that targeted inhibition of miR-182 may prevent leptomeningeal spread in patients with non-SHH-MB.

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Acknowledgments

We gratefully thank the patients and families for participating in this research. Jennifer Diemer and Anna Schöttler are acknowledged for excellent technical assistance. Support by the DKFZ Light Microscopy Facility is gratefully acknowledged. This study was supported by a grant from the Deutsche Kinderkrebsstiftung and from the “Tumorzentrum Heidelberg” to S.M.P.; a “Young Investigator Fellowship” of the Medical Faculty of Heidelberg and financial support by the Eliteprogramme for Postdocs (Baden-Württemberg Stiftung) to M.R.

Conflict of interest

The authors declare no conflicts of interest.

Author information

Correspondence to Stefan M. Pfister.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary Video 1 DAOY cells stably transfected with miR-182 were seeded closely to the channels. The third channel on the right hand side, a representative permeation activity was observed in this channel. Duration 24 h; scale bar, 100μm (MPG 2286 kb)

Supplementary Video 2 DAOY cells stably transfected with empty vector (pCMX). All four channels on the right hand side showed the typical penetrative behavior. These cells continuously tried to enter the channels within the whole experimental period, while nuclei are not able to enter the channels. Duration 24 h; scale bar, 100μm (MPG 2194 kb)

Supplementary Video 3 DAOY cells with miR-182 knockdown after 24h. Less invasive activity was observed and the cells were only trying to enter the channels for short periods of time. Duration 24 h; scale bar, 100μm (MPG 2582 kb)

Supplementary Video 4 DAOY cells with scrambled siRNA treatment. A DAOY cell invaded into the channel and migrated until the middle of the channel. Then the cell returned to the starting point. This special invasion behavior could be found in the second channel on right hand side. Duration 24 h; scale bar, 100μm (MPG 2414 kb)

Supplementary Video 5 Med8A cells stably transfected with miR-182. Most of the cells remained in the same position, and generally did not enter the channel. The fourth channel on the left side showed a rare example for a single invasion. Duration 24 h; scale bar, 100μm (MPG 2128 kb)

Supplementary Video 6 Med8A cells stably transfected with empty vector. All cells included in the field of view remained in a similar position. No invasive behaviors observed at all. Duration 23:20 h; scale bar, 100μm (MPG 2456 kb)

Supplementary Video 1 DAOY cells stably transfected with miR-182 were seeded closely to the channels. The third channel on the right hand side, a representative permeation activity was observed in this channel. Duration 24 h; scale bar, 100μm (MPG 2286 kb)

Supplementary Video 2 DAOY cells stably transfected with empty vector (pCMX). All four channels on the right hand side showed the typical penetrative behavior. These cells continuously tried to enter the channels within the whole experimental period, while nuclei are not able to enter the channels. Duration 24 h; scale bar, 100μm (MPG 2194 kb)

Supplementary Video 3 DAOY cells with miR-182 knockdown after 24h. Less invasive activity was observed and the cells were only trying to enter the channels for short periods of time. Duration 24 h; scale bar, 100μm (MPG 2582 kb)

Supplementary Video 4 DAOY cells with scrambled siRNA treatment. A DAOY cell invaded into the channel and migrated until the middle of the channel. Then the cell returned to the starting point. This special invasion behavior could be found in the second channel on right hand side. Duration 24 h; scale bar, 100μm (MPG 2414 kb)

Supplementary Video 5 Med8A cells stably transfected with miR-182. Most of the cells remained in the same position, and generally did not enter the channel. The fourth channel on the left side showed a rare example for a single invasion. Duration 24 h; scale bar, 100μm (MPG 2128 kb)

Supplementary Video 6 Med8A cells stably transfected with empty vector. All cells included in the field of view remained in a similar position. No invasive behaviors observed at all. Duration 23:20 h; scale bar, 100μm (MPG 2456 kb)

Supplementary material 7 (DOC 158 kb)

Supplementary material 8 (PPT 6229 kb)

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Bai, A.H.C., Milde, T., Remke, M. et al. MicroRNA-182 promotes leptomeningeal spread of non-sonic hedgehog-medulloblastoma. Acta Neuropathol 123, 529–538 (2012). https://doi.org/10.1007/s00401-011-0924-x

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Keywords

  • hsa-miR-182
  • Metastatic dissemination
  • SHH pathway
  • Medulloblastoma