Targeting neuroblastoma stem cells with retinoic acid and proteasome inhibitor

PLoS One. 2013 Oct 7;8(10):e76761. doi: 10.1371/journal.pone.0076761. eCollection 2013.

Abstract

Background: Neuroblastma cell lines contain a side-population of cells which express stemness markers. These stem-like cells may represent the potential underlying mechanism for resistance to conventional therapy and recurrence of neuroblastoma in patients.

Methodology/principal findings: To develop novel strategies for targeting the side-population of neurobastomas, we analyzed the effects of 13-cis-retinoic acid (RA) combined with the proteasome inhibitor MG132. The short-term action of the treatment was compared with effects after a 5-day recovery period during which both chemicals were withdrawn. RA induced growth arrest and differentiation of SH-SY5Y and SK-N-BE(2) neuroblastoma cell lines. Inhibition of the proteasome caused apoptosis in both cell lines, thus, revealing the critical role of this pathway in the regulated degradation of proteins involved in neuroblastoma proliferation and survival. The combination of RA with MG132 induced apoptosis in a dose-dependent manner, in addition to promoting G2/M arrest in treated cultures. Interestingly, expression of stem cell markers such as Nestin, Sox2, and Oct4 were reduced after the recovery period of combined treatment as compared with untreated cells or treated cells with either compound alone. Consistent with this, neurosphere formation was significantly impaired by the combined treatment of RA and MG132.

Conclusions: Given that stem-like cells are associated with resistant to conventional therapy and are thought to be responsible for relapse, our results suggest that dual therapy of RA and proteasome inhibitor might be beneficial for targeting the side-population of cells associated residual disease in high-risk neuroblastoma.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / drug effects
  • Blotting, Western
  • Cell Differentiation / drug effects
  • Cell Line, Tumor
  • Cell Proliferation / drug effects
  • Dose-Response Relationship, Drug
  • Drug Synergism
  • Flow Cytometry
  • G2 Phase Cell Cycle Checkpoints / drug effects
  • Humans
  • Leupeptins / pharmacology*
  • Microscopy, Confocal
  • Neoplastic Stem Cells / drug effects*
  • Neoplastic Stem Cells / metabolism
  • Neoplastic Stem Cells / pathology
  • Nestin / metabolism
  • Neuroblastoma / metabolism
  • Neuroblastoma / pathology
  • Octamer Transcription Factor-3 / metabolism
  • Proteasome Inhibitors / pharmacology*
  • SOXB1 Transcription Factors / metabolism
  • Side-Population Cells / drug effects
  • Side-Population Cells / metabolism
  • Side-Population Cells / pathology
  • Time Factors
  • Tretinoin / pharmacology*

Substances

  • Leupeptins
  • NES protein, human
  • Nestin
  • Octamer Transcription Factor-3
  • POU5F1 protein, human
  • Proteasome Inhibitors
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • Tretinoin
  • benzyloxycarbonylleucyl-leucyl-leucine aldehyde

Grants and funding

This work was supported by SAF2009-08334 and SAF2012-37330 grants from Spanish Ministry of Economy and Competitivity and NeuroblastoVal from Gent per Gent. A part of this work was also funded by the Regenerative Medicine program of the Valencian Community (ISCIII). BH was supported by a post-doctoral fellowship from Bancaixa Foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.