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Inhibition of autophagy significantly increases the antitumor effect of Abiraterone in prostate cancer

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Abstract

Purpose

Abiraterone acetate (AA) plus prednisone is an approved treatment of advanced prostate cancer (PCa). Autophagy is linked to drug resistance in numerous types of cancers. We hypothesized, that upregulation of autophagy is one of the mechanisms by which PCa cells survive AA anti-tumor treatment and therefore evaluated the potential effect of a combination with autophagy inhibition.

Methods

Human PCa LNCaP cell lines were cultured in steroid-free medium and treated with AA. Autophagy was inhibited by 3-methyladenine, chloroquine and ATG5 siRNA knock-down. Cell viability and apoptosis was assessed by flow cytometry and fluorescence microscopy, and autophagy was monitored by immunohistochemistry, AUTOdot and Western blotting.

Results

Western blot revealed upregulation of ATG5 and LC3 II with a reduction of p62 protein expression in AA-treated cells, indicating upregulation of autophagy. These data were supported by results obtained with immunocytochemistry and AUTOdot assays. Using flow cytometry, we showed that combining AA with autophagy inhibition significantly impaired cell viability (1.3–1.6-fold, p < 0.001) and increased apoptosis (1.4–1.5-fold, p < 0.001) compared to AA treatment alone.

Conclusions

AA activates autophagy as a cytoprotective mechanism in LNCaP prostate cancer cells and targeting of autophagy enhances the antitumor effect of the compound.

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Acknowledgements

This study was supported by Janssen Pharmaceutica NV and Fond zur Förderung des akademischen Nachwuchses (FAN). Special thanks to Damina Balmer for the critical assessment of this manuscript.

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Author notes

  1. Ashkan Mortezavi and Souzan Salemi have contributed equally to this work.

Authors and Affiliations

  1. Laboratory for Tissue Engineering and Stem Cell Therapy, Department of Urology, University Hospital Zürich, Frauenklinikstrasse 10, 8091, Zurich, Switzerland

    Ashkan Mortezavi, Souzan Salemi, Benedikt Kranzbühler, Oliver Gross, Tullio Sulser & Daniel Eberli

  2. Institute of Pharmacology, University of Bern, Inselspital, INO-F, 3010, Bern, Switzerland

    Hans-Uwe Simon

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  1. Ashkan Mortezavi
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Contributions

AM and SS contributed equally to this work: project development, carried out all the experiments, data collection, interpretation and manuscript writing. BK and OG: data collection and revising the manuscript. TS: revising the manuscript. HUS: data interpretation and revising the manuscript and DE: project development, data interpretation and manuscript writing.

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Correspondence to Daniel Eberli.

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The authors declare that they have no conflict of interest.

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

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Informed consent was obtained from all the individual participants included in the study.

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Supplementary material 1 (DOCX 38 kb)

345_2018_2385_MOESM2_ESM.tif

Fig. 1S: A-E: A: WST cell proliferation assay: PC3 cells treated with AA 10 µM, 3MA 5 mM, Chl 20 µM and combinations for 2 and 4 days. Absorbance was measured daily at 450 nm. Bars represent mean ± SEM (n = 3). B: Cell death assay. PC3 cells were cultured in the presence and absence of AA 10 µM, 3MA 5 mM, Chl 20 µM and in combination of AA and 3MA and AA and Chl for 4 days. Cell viability was assessed using ethidium bromide and measured by FACS. Values are mean ± SEM of three independent experiments. C: Immunoblotting analysis (day 4) of PC3 cells. No changes of 55 kDa ATG5-ATG12 expression indicating no autophagy activation in response to different treatments. Supplementary material 2 (TIFF 7906 kb)

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Mortezavi, A., Salemi, S., Kranzbühler, B. et al. Inhibition of autophagy significantly increases the antitumor effect of Abiraterone in prostate cancer. World J Urol 37, 351–358 (2019). https://doi.org/10.1007/s00345-018-2385-5

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  • DOI: https://doi.org/10.1007/s00345-018-2385-5

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