Abrogation of Autophagy by Chloroquine Alone or in Combination with mTOR Inhibitors Induces Apoptosis in Neuroendocrine Tumor Cells

Shani Avniel-Polak, Gil Leibowitz, Yael Riahi, Benjamin Glaser, David J. Gross, Simona Grozinsky-Glasberg

Research output: Contribution to journalArticlepeer-review

20 Scopus citations

Abstract

Background: Everolimus (RAD001), an mTORC1 inhibitor, demonstrated promising, but limited, anticancer effects in neuroendocrine tumors (NETs). Torin1 (a global mTOR inhibitor) and NVP-BEZ235 (a PI3K/mTOR inhibitor) seem to be more effective than RAD001. Autophagy, a degradation pathway that may promote tumor growth, is regulated by mTOR; mTOR inhibition results in stimulation of autophagy. Chloroquine (CQ) inhibits autophagy. Aim: To explore the effect of CQ alone or in combination with RAD001, Torin1 or NVP-BEZ235 on autophagy and on NET cell viability, proliferation and apoptosis. Methods: The NET cell line BON1 was treated with CQ with or without different mTOR inhibitors. siRNA against ATG5/7 was used to genetically inhibit autophagy. Cellular viability was examined by XTT, proliferation by Ki-67 staining and cell cycles by flow cytometry. Apoptosis was analyzed by Western blotting for cleaved caspase 3 and staining for annexin V; autophagy was evaluated by Western blotting and immunostaining for LC3. Results: RAD001, Torin1, NVP-BEZ235 and CQ all decreased BON1 cell viability. The effect of RAD001 was smaller than that of the other mTOR inhibitors or CQ. Torin1 and NVP-BEZ235 markedly inhibited cell proliferation, without inducing apoptosis. CQ similarly decreased cell proliferation, while robustly increasing apoptosis. Treatment with Torin1 or NVP-BEZ235 together with CQ was additive on viability, without increasing CQ-induced apoptosis. Inhibition of autophagy by ATG5/7 knockdown increased apoptosis in the presence or absence of mTOR inhibitors, mimicking the CQ effects. Conclusion: CQ inhibits NET growth by inducing apoptosis and by inhibiting cell proliferation, probably via inhibition of autophagy. CQ may potentiate the antitumor effect of mTOR inhibitors.

Original languageAmerican English
Pages (from-to)724-737
Number of pages14
JournalNeuroendocrinology
Volume103
Issue number6
DOIs
StatePublished - 1 Sep 2016
Externally publishedYes

Bibliographical note

Publisher Copyright:
© 2015 S. Karger AG, Basel. Copyright: All rights reserved.

Keywords

  • Autophagy
  • Chloroquine
  • Neuroendocrine cells
  • mTOR inhibitors

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