Journal of Neuro-Oncology

, Volume 134, Issue 2, pp 259–268 | Cite as

Synergistic inhibition of glioma cell proliferation by Withaferin A and tumor treating fields

  • Edwin Chang
  • Christoph Pohling
  • Nooshin Beygui
  • Chirag B. Patel
  • Jarrett Rosenberg
  • Dong Ho Ha
  • Sanjiv S. Gambhir
Laboratory Investigation


Glioblastoma (GBM) is the most aggressive and lethal form of brain cancer. Standard therapies are non-specific and often of limited effectiveness; thus, efforts are underway to uncover novel, unorthodox therapies against GBM. In previous studies, we investigated Withaferin A, a steroidal lactone from Ayurvedic medicine that inhibits proliferation in cancers including GBM. Another novel approach, tumor treating fields (TTFields), is thought to disrupt mitotic spindle formation and stymie proliferation of actively dividing cells. We hypothesized that combining TTFields with Withaferin A would synergistically inhibit proliferation in glioblastoma. Human glioblastoma cells (GBM2, GBM39, U87-MG) and human breast adenocarcinoma cells (MDA-MB-231) were isolated from primary tumors. The glioma cell lines were genetically engineered to express firefly luciferase. Proliferative potential was assessed either by bioluminescence imaging or cell counting via hemocytometer. TTFields (4 V/cm) significantly inhibited growth of the four cancer cell lines tested (n = 3 experiments per time point, four measurements per sample, p < 0.02 at least; 2-way ANOVA, control vs. treatment). The combination of Withaferin A (10–100 nM) with TTFields significantly inhibited the growth of the glioma cells to a degree beyond that of Withaferin A or TTFields alone. The interaction of the Withaferin A and TTFields on glioma cells was found to be synergistic in nature (p < 0.01, n = 3 experiments). These findings were validated by both bioluminescence and hemocytometric measurements. The combination of Withaferin A with TTFields represents a novel approach to treat GBM in a manner that is likely better than either treatment alone and that is synergistic.


Alternating electric fields Combination therapy Glioblastoma Synergy Tumor treating fields Withaferin A 



Analysis of variance


Bioluminescence imaging




Dulbecco’s modified eagle’s medium


Human epidermal growth factor


Epidermal growth factor receptor


Epidermal growth factor receptor variant III


Fetal bovine serum


Food and drug administration


Human fibroblast growth factor




Patient-derived glioblastoma cell culture (from Stanford University School of Medicine)


GBM2 that was genetically modified to express a fusion protein of firefly luciferase and GFP


Patient-derived glioblastoma cell culture (from University of California at San Diego School of Medicine)


GBM39 that was genetically modified to express firefly luciferase


Green fluorescent protein


Fusion protein of GFP and firefly luciferase


Human breast adenocarcinoma cancer cell line (from ATCC)


Phosphate-buffered saline


Human platelet-derived growth factor variant AA


Human platelet-derived growth factor variant BB


Tumor treating field


Human-derived GBM cell line that was purchased from ATCC


U87-MG that was genetically modified to express a fusion protein of firefly luciferase and GFP

Supplementary material

11060_2017_2534_MOESM1_ESM.docx (24 kb)
Supplementary material 1 (DOCX 24 KB)
11060_2017_2534_MOESM2_ESM.docx (17 kb)
Supplementary material 2 (DOCX 17 KB)
11060_2017_2534_MOESM3_ESM.pptx (4.5 mb)
Supplementary material 3 (PPTX 4589 KB)


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Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  1. 1.Department of Radiology, Molecular Imaging Program at Stanford, Canary Center for Early Cancer DetectionStanford UniversityPalo AltoUSA
  2. 2.Scripps CollegeClaremontUSA
  3. 3.Department of RadiologyDong-A University Medical CenterBusanKorea

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