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Ganoderic acid A/DM-induced NDRG2 over-expression suppresses high-grade meningioma growth

  • A. DasEmail author
  • M. Alshareef
  • F. HendersonJr.
  • J. L. Martinez Santos
  • W. A. VandergriftIII
  • S. M. Lindhorst
  • A. K. Varma
  • L. Infinger
  • S. J. Patel
  • D. Cachia
Research Article

Abstract

Purpose

N-myc downstream-regulated gene 2 (NDRG2) is down-regulated in grade-III meningioma [anaplastic meningioma (AM)] and associated with clinically aggressive behavior. Current therapies in the treatment of high-grade meningioma are lacking with limited success. This study aims to validate the effect of NDRG2-targeted therapy using structurally related bioactive triterpene compounds derived from the edible mushroom Ganoderma lucidum (ganoderic acid A:GA-A/ganoderic acid DM:GA-DM) in human AM in relevant pre-clinical models.

Methods

Tissue samples from the AM tumor regions of three human patients and control non-tumor samples were used to analyze the expression pattern of NDRG2. In vitro cell culture and in vivo cell-line-derived orthotopic xenograft animal models of AM were utilized to assess efficacy of treatment with GA-A/DM.

Results

Downregulation of NDRG2 expression was observed in surgically resected high-grade meningiomas compared to normal brain. These results prompt us to use NDRG2-targeting agents GA-A/DM. In vitro results showed that 72-h treatments of 25 µM GA-A/DM induced AM cell death, upregulate NDRG2 protein expression, downregulate NDRG2 promoter methylation in meningioma cells as compared to azacitidine and decitabine, the most commonly used demethylating agents. Our results also demonstrated that GA-A/DM does not have any detrimental effect on normal human neurons and arachnoid cells. GA-A/DM promoted apoptotic factors (Bax) while suppressing MMP-9, p-P13K, p-AKT, p-mTOR, and Wnt-2 protein expression. RNAi-mediated knockdown of NDRG2 protein expression increased tumor proliferation, while forced expression of wt-NDRG2 decreased proliferation in an in vitro model. Magnetic resonance (MR) imaging and Hematoxylin (H&E) staining demonstrated gross reduction of tumor volume in GA-A/DM treated mice at 5 weeks when compared with saline-treated orthotopic AM xenografted controls. There was an overall decrease in tumor cell proliferation with increased survival in GA-A/DM-treated animals. Enzyme assays showed that GA-A/DM did not negatively impact hepatic function.

Conclusion

GA-A/DM may be a promising natural therapeutic reagent in the treatment of AM by suppressing growth via NDRG2 modulation and altering of intracellular signal pathways. We have shown it could potentially be an effective treatment for AM with decreased cellular proliferation in vitro, decreased tumor volume and increased survival in vivo.

Keywords

Ganoderic acid A/DM Anaplastic meningioma (AM) NDRG2 Animal model 

Notes

Author contributions

Conceived and designed the experiments: AD and DC. Analyzed the data: AD, MA, and DC. Wrote the first draft of the manuscript: AD and DC. Contributed to the writing of the manuscript: AD, MA, FH, SJP, SML, LI, AKV, WAV, and JLMS (all authors). Agree with manuscript results and conclusions: all authors. Jointly developed the structure and arguments for the paper: all authors. Made critical revisions and approved final version: AD and DC. All authors reviewed and approved of the final manuscript.

Funding

Completion of this project was made possible in part by the MUSC Neuroscience Institute Pilot Grants (2017-18), and Funds from the Department of Neurosurgery.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.

Informed consent

Subjects gave their written, informed consent to participate.

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Copyright information

© Federación de Sociedades Españolas de Oncología (FESEO) 2019

Authors and Affiliations

  • A. Das
    • 1
    Email author
  • M. Alshareef
    • 1
  • F. HendersonJr.
    • 1
  • J. L. Martinez Santos
    • 1
  • W. A. VandergriftIII
    • 1
  • S. M. Lindhorst
    • 1
  • A. K. Varma
    • 1
  • L. Infinger
    • 1
  • S. J. Patel
    • 1
  • D. Cachia
    • 1
  1. 1.Department of Neurosurgery (Divisions of Neuro-oncology) and MUSC Brain and Spine Tumor Program CSB 310Medical University of South CarolinaCharlestonUSA

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