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Effects of sequentially applied single and combined temozolomide, hydroxychloroquine and AT101 treatment in a long-term stimulation glioblastoma in vitro model

  • Original Article – Cancer Research
  • Published:
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Abstract

Purpose

Glioblastoma multiforme (GBM) is a poorly curable disease due to its heterogeneity that enables single cells to survive treatment regimen and initiate tumor regrowth. Although some progress in therapy has been achieved in the last years, the efficient treatment of GBMs is still a clinical challenge. Besides the standard therapeutic drug temozolomide (TMZ), quinoline-based antimalarial drugs such as hydroxychloroquine (HCQ) and BH3 mimetics such as AT101 were considered as possible drugs for GBM therapy.

Methods

We investigated the effects of sequentially applied single and combined TMZ, HCQ and AT101 treatments in a long-term stimulation GBM in vitro model. We performed all investigations in parallel in human astrocytes and two differentially TMZ-responsive human GBM cell lines and adjusted used drug concentrations to known liquor/plasma concentrations in patients. We determined amounts of dead cells and still remaining growth rates and depicted our results in a heatmap-like summary to visualize which sequential long-term treatment schedule seemed to be most promising.

Results

We showed that sequential stimulations yielded higher cytotoxicity and better tumor growth control in comparison to single TMZ treatment. This was especially the case for the sequences TMZ/HCQ and TMZ + AT101/AT101 which was as effective as the non-sequential combination TMZ + AT101. Importantly, those affected both less and more TMZ-responsive glioma cell lines, whilst being less harmful for astrocytes in comparison to single TMZ treatment.

Conclusions

Sequential treatment with mechanistically different acting drugs might be an option to reduce side effects in long-term treatment, for example in local administration approaches.

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Abbreviations

CHOP:

CCAAT-enhancer-binding protein homologous protein

CI:

Combination index

CQ:

Chloroquine

DMEM:

Dulbecco’s modified Eagle’s medium

DMSO:

Dimethyl sulfoxide

ER:

Endoplasmatic reticulum

FBS:

Fetal bovine serum

GAAD-153:

Growth arrest and DNA-damage-inducible protein

GBM:

Glioblastoma multiforme

HCQ:

Hydroxychloroquine

PARP:

Poly-ADP ribose polymerase

TMZ:

Temozolomide

QNX:

Quinacrine

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Acknowledgements

We thank Fereshteh Ebrahim, Brigitte Rehmke, Judith Becker and Sonja Dahle for expert technical assistance.

Funding

This study was funded by the German Research Foundation (DFG) as part as of the Research Training Group “Materials4Brain” (GRK2154; P3, P7 and P8).

Author information

Author notes

  1. Vivian Adamski and Christina Schmitt share first authorship.

  2. Kirsten Hattermann and Janka Held-Feindt share senior authorship.

Authors and Affiliations

  1. Department of Neurosurgery, University Medical Center Schleswig-Holstein UKSH, Campus Kiel, Arnold-Heller-Str.3, Building 41, 24105, Kiel, Germany

    Vivian Adamski, Michael Synowitz & Janka Held-Feindt

  2. Department of Anatomy, University of Kiel, 24118, Kiel, Germany

    Christina Schmitt, Ralph Lucius & Kirsten Hattermann

  3. Institute for Materials Science, University of Kiel, 24143, Kiel, Germany

    Florian Ceynowa & Rainer Adelung

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  1. Vivian Adamski
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Contributions

JHF, KH conceived and designed the study; VA, CS, JHF and KH performed the experiments and analyzed the data; FC, RA, RL and MS contributed materials and assisted in data analysis; JHF and KH wrote the paper, and all authors revised the manuscript.

Corresponding author

Correspondence to Janka Held-Feindt.

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Conflict of interest

Author Vivian Adamski declares that she has no conflict of interest. Author Christina Schmitt declares that she has no conflict of interest. Author Florian Ceynowa declares that he has no conflict of interest. Author Rainer Adelung declares that he has no conflict of interest. Author Ralph Lucius declares that he has no conflict of interest. Author Michael Synowitz declares that he has no conflict of interest. Author Kirsten Hattermann declares that she has no conflict of interest. Author Janka Held-Feindt declares that she has no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Electronic supplementary material

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432_2018_2680_MOESM1_ESM.tif

Supplementary material 1 Figure 1 Supplement : Treatment schedules for the long-term GBM in vitro model. A. Cells were treated for 3 days with the gold standard therapeutic temozolomide (TMZ), switching on day 3 either to the combined treatment of TMZ plus an alternative drug [shown as TMZ+AT101 or TMZ+hydroxychloroquine(HCQ)], or to the single alternative drug (AT101 or HCQ), respective staying with the TMZ treatment. B. Cells were treated for 3 days with one of the single alternative drugs (AT101 or HCQ), switching on day 3 to either the combined therapy with TMZ, to the gold standard therapeutic TMZ or staying with the single alternative drug. C. Cells were treated for 3 days with the combined treatment of TMZ plus one alternative drug, switching on day 3 to either one of the single drugs TMZ, AT101 or HCQ respective staying with the combined therapy until day 6 (TIF 676 KB)

432_2018_2680_MOESM2_ESM.tif

Supplementary material 2 Figure 2 Supplement : Effect of selected (non)-sequential treatments on growth rates of astrocytes and two different GBM cell lines treated according to the treatment schedules (ref. Fig. 1 Supplement). The relative growth rates for day 3 and day 6 of SVGA (A), A172 (B) and LN229 (C) were determined compared to the number of cells at day 0 of the treatment. Growth rates of cell treated with the gold standard TMZ are highlighted with dashed lines. Means are significantly different for *p<0.05, **p<0.01 respective ***p<0.001 (TIF 1339 KB)

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Adamski, V., Schmitt, C., Ceynowa, F. et al. Effects of sequentially applied single and combined temozolomide, hydroxychloroquine and AT101 treatment in a long-term stimulation glioblastoma in vitro model. J Cancer Res Clin Oncol 144, 1475–1485 (2018). https://doi.org/10.1007/s00432-018-2680-y

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  • DOI: https://doi.org/10.1007/s00432-018-2680-y

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