Temozolomide induces activation of Wnt/β-catenin signaling in glioma cells via PI3K/Akt pathway: implications in glioma therapy

Abstract

Glioblastoma (GBM) is the most aggressive type of glioma. Temozolomide (TMZ) is currently the drug of choice used for post-operative chemotherapy of GBM. However, the presence of intrinsic and acquired resistance hinders the success of chemotherapy. To understand the TMZ resistant mechanisms in glioma, we investigated the alterations in cellular signaling pathways by performing transcriptome analysis of TMZ treated glioma cells. Gene Set Enrichment Analysis (GSEA) indicated a significant enrichment of Wnt/β-catenin signaling besides many other pathways in TMZ treated cells. Further, we demonstrate that TMZ treatment increased the activity from TOPflash reporter, (a Wnt responsive reporter), enhanced the levels of pGSK-3β (S9) and reduced the levels of p-β-catenin (S33/37/T41) with a concomitant increase in transcript and protein levels of Wnt targets in a concentration and time-dependent manner. While TMZ treated cells did not show alteration in any of the Wnt ligands, PI3K inhibitor (LY294002) treatment repressed Akt activation and abolished the TMZ–mediated induction of Wnt/β-catenin pathway. In addition, we show that Wnt/β-catenin signaling activation by TMZ is independent of ATM/Chk2 pathway. Further, we also demonstrate the activation of mTOR pathway after TMZ treatment. Thus, our results demonstrate that activation of Wnt/β-catenin pathway involves an ATM/Chk2- independent PI3K/Akt/GSK-3 cascade in TMZ treated cells and further provides mechanistic basis for the chemoresistance of glioma to TMZ.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2

Abbreviations

GBM:

Glioblastoma

TMZ:

Temozolomide

DAVID:

Database for Annotation, Visualization and Integrated Discovery

GSEA:

Gene Set Enrichment Analysis

MSigDB:

Molecular signature database

References

  1. Bezler M, Hengstler JG, Ullrich A. Inhibition of doxorubicin-induced HER3-PI3K-AKT signalling enhances apoptosis of ovarian cancer cells. Mol Oncol. 2012;6:516–29.

    CAS  Article  PubMed Central  Google Scholar 

  2. Bhanot P, Brink M, Samos CH, Hsieh JC, Wang Y, Macke JP, et al. A new member of the frizzled family from Drosophila functions as a wingless receptor. Nature. 1996;382:225–30.

    CAS  Article  PubMed Central  Google Scholar 

  3. Galli C, Piemontese M, Lumetti S, Manfredi E, Macaluso GM, Passeri G. GSK3b-inhibitor lithium chloride enhances activation of Wnt canonical signaling and osteoblast differentiation on hydrophilic titanium surfaces. Clin Oral Implants Res. 2013;24:921–7.

    CAS  Article  PubMed Central  Google Scholar 

  4. Holohan C, Van Schaeybroeck S, Longley DB, Johnston PG. Cancer drug resistance: an evolving paradigm. Nat Rev Cancer. 2013;13:714–26.

    CAS  Article  PubMed Central  Google Scholar 

  5. Lin F, de Gooijer MC, Hanekamp D, Chandrasekaran G, Buil LC, Thota N, et al. PI3K-mTOR pathway inhibition exhibits efficacy against high-grade glioma in clinically relevant mouse models. Clin Cancer Res. 2017;23:1286–98.

    CAS  Article  PubMed Central  Google Scholar 

  6. Mootha VK, Lindgren CM, Eriksson KF, Subramanian A, Sihag S, Lehar J, et al. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet. 2003;34:267–73.

    CAS  Article  Google Scholar 

  7. Niida A, Hiroko T, Kasai M, Furukawa Y, Nakamura Y, Suzuki Y, et al. DKK1, a negative regulator of Wnt signaling, is a target of the beta-catenin/TCF pathway. Oncogene. 2004;23:8520–6.

    CAS  Article  PubMed Central  Google Scholar 

  8. Pessina S, Cantini G, Kapetis D, Cazzato E, Di Ianni N, Finocchiaro G, et al. The multidrug-resistance transporter Abcc3 protects NK cells from chemotherapy in a murine model of malignant glioma. Oncoimmunology. 2016;5:e1108513.

    Article  PubMed Central  Google Scholar 

  9. Rheinbay E, Suva ML, Gillespie SM, Wakimoto H, Patel AP, Shahid M, et al. An aberrant transcription factor network essential for Wnt signaling and stem cell maintenance in glioblastoma. Cell Rep. 2013;3:1567–79.

    CAS  Article  PubMed Central  Google Scholar 

  10. Riganti C, Salaroglio IC, Caldera V, Campia I, Kopecka J, Mellai M, et al. Temozolomide downregulates P-glycoprotein expression in glioblastoma stem cells by interfering with the Wnt3a/glycogen synthase-3 kinase/beta-catenin pathway. Neuro Oncol. 2013;15:1502–17.

    CAS  Article  PubMed Central  Google Scholar 

  11. Shen D, Guo CC, Wang J, Qiu ZK, Sai K, Yang QY, et al. Interferon-alpha/beta enhances temozolomide activity against MGMT-positive glioma stem-like cells. Oncol Rep. 2015;34:2715–21.

    CAS  Article  PubMed Central  Google Scholar 

  12. Stephens L, Anderson K, Stokoe D, Erdjument-Bromage H, Painter GF, Holmes AB, et al. Protein kinase B kinases that mediate phosphatidylinositol 3,4,5-trisphosphate-dependent activation of protein kinase B. Science. 1998;279:710–4.

    CAS  Article  PubMed Central  Google Scholar 

  13. Stupp R, Hegi ME, Mason WP, van den Bent MJ, Taphoorn MJ, Janzer RC, et al. Effects of radiotherapy with concomitant and adjuvant temozolomide versus radiotherapy alone on survival in glioblastoma in a randomised phase III study: 5-year analysis of the EORTC-NCIC trial. Lancet Oncol. 2009;10:459–66.

    CAS  Article  Google Scholar 

  14. Stupp R, Mason WP, van den Bent MJ, Weller M, Fisher B, Taphoorn MJ, et al. Radiotherapy plus concomitant and adjuvant temozolomide for glioblastoma. N Engl J Med. 2005;352:987–96.

    CAS  Article  Google Scholar 

  15. Subramanian A, Tamayo P, Mootha VK, Mukherjee S, Ebert BL, Gillette MA, et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc Natl Acad Sci U S A. 2005;102:15545–50.

    CAS  Article  PubMed Central  Google Scholar 

  16. Tan Z, Song L, Wu W, Zhou Y, Zhu J, Wu G, et al. TRIM14 promotes chemoresistance in gliomas by activating Wnt/beta-catenin signaling via stabilizing Dvl2. Oncogene. 2018;37:5403–15.

    CAS  Article  PubMed Central  Google Scholar 

  17. Ulasov IV, Nandi S, Dey M, Sonabend AM, Lesniak MS. Inhibition of Sonic hedgehog and Notch pathways enhances sensitivity of CD133(+) glioma stem cells to temozolomide therapy. Mol Med. 2011;17:103–12.

    CAS  Article  PubMed Central  Google Scholar 

  18. Xu N, Lao Y, Zhang Y, Gillespie DA. Akt: a double-edged sword in cell proliferation and genome stability. J Oncol. 2012;2012:951724.

    Article  PubMed Central  Google Scholar 

  19. Zhang LH, Yin AA, Cheng JX, Huang HY, Li XM, Zhang YQ, et al. TRIM24 promotes glioma progression and enhances chemoresistance through activation of the PI3K/Akt signaling pathway. Oncogene. 2015;34:600–10.

    CAS  Article  PubMed Central  Google Scholar 

Download references

Acknowledgments

We thank Dr. Subba Rao and Dr. Deepak Saini (I.I.Sc., Bangalore) for providing the shRNA constructs. We acknowledge use of GSEA software and MSigDB (Mootha et al. 2003; Subramanian et al. 2005).

Funding

VS acknowledge IISc for the research fellowship. KS acknowledges CSIR, DST, SERB, CEFIPRA, and DBT, Government of India, for research grants. Infrastructure support by funding from DST-FIST and UGC (Center for Advanced Studies in Molecular Microbiology) to MCB and DBT-IISc partnership program is acknowledged. KS is a J. C. Bose Fellow of the Department of Science and Technology.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Kumaravel Somasundaram.

Ethics declarations

Conflict of interest

The authors declare no conflict of interest.

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

ESM1

(PPTX 238 kb)

ESM2

(PPTX 10.1 mb)

ESM3

(PPTX 88.1 kb)

ESM4

(PPTX 134 kb)

ESM5

(PPTX 324 kb)

ESM6

(PPTX 257 kb)

ESM7

(PPTX 197 kb)

ESM8

(DOCX 24.8 kb)

ESM9

(XLS 862 kb)

ESM10

(XLS 91.5 kb)

ESM11

(XLS 27.0 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Tomar, V.S., Patil, V. & Somasundaram, K. Temozolomide induces activation of Wnt/β-catenin signaling in glioma cells via PI3K/Akt pathway: implications in glioma therapy. Cell Biol Toxicol 36, 273–278 (2020). https://doi.org/10.1007/s10565-019-09502-7

Download citation

Keywords

  • Glioblastoma
  • Microarray
  • Temozolomide
  • Transcriptome
  • Resistance