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Screen for chemical modulators of autophagy reveals novel therapeutic inhibitors of mTORC1 signaling

Screen for chemical modulators of autophagy reveals novel therapeutic inhibitors of mTORC1 signaling
Screen for chemical modulators of autophagy reveals novel therapeutic inhibitors of mTORC1 signaling
Background
Mammalian target of rapamycin complex 1 (mTORC1) is a protein kinase that relays nutrient availability signals to control numerous cellular functions including autophagy, a process of cellular self-eating activated by nutrient depletion. Addressing the therapeutic potential of modulating mTORC1 signaling and autophagy in human disease requires active chemicals with pharmacologically desirable properties.

Methodology/Principal Findings
Using an automated cell-based assay, we screened a collection of >3,500 chemicals and identified three approved drugs (perhexiline, niclosamide, amiodarone) and one pharmacological reagent (rottlerin) capable of rapidly increasing autophagosome content. Biochemical assays showed that the four compounds stimulate autophagy and inhibit mTORC1 signaling in cells maintained in nutrient-rich conditions. The compounds did not inhibit mTORC2, which also contains mTOR as a catalytic subunit, suggesting that they do not inhibit mTOR catalytic activity but rather inhibit signaling to mTORC1. mTORC1 inhibition and autophagosome accumulation induced by perhexiline, niclosamide or rottlerin were rapidly reversed upon drug withdrawal whereas amiodarone inhibited mTORC1 essentially irreversibly. TSC2, a negative regulator of mTORC1, was required for inhibition of mTORC1 signaling by rottlerin but not for mTORC1 inhibition by perhexiline, niclosamide and amiodarone. Transient exposure of immortalized mouse embryo fibroblasts to these drugs was not toxic in nutrient-rich conditions but led to rapid cell death by apoptosis in starvation conditions, by a mechanism determined in large part by the tuberous sclerosis complex protein TSC2, an upstream regulator of mTORC1. By contrast, transient exposure to the mTORC1 inhibitor rapamycin caused essentially irreversible mTORC1 inhibition, sustained inhibition of cell growth and no selective cell killing in starvation.

Conclusion/Significance
The observation that drugs already approved for human use can reversibly inhibit mTORC1 and stimulate autophagy should greatly facilitate the preclinical and clinical testing of mTORC1 inhibition for indications such as tuberous sclerosis, diabetes, cardiovascular disease and cancer.
1932-6203
e7124
Balgi, Aruna D.
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Fonseca, Bruno D.
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Donohue, Elizabeth
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Tsang, Trevor C. F.
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Lajoie, Patrick
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Proud, Christopher G.
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Nabi, Ivan R.
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Roberge, Michel
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Balgi, Aruna D.
70f66417-efe1-494f-a8ea-730b5f1ebc29
Fonseca, Bruno D.
bd81f9c7-365f-4636-a043-d204a8c4ccb9
Donohue, Elizabeth
e11f5793-a077-4c33-86b0-a1a82ed698be
Tsang, Trevor C. F.
d9e4cefa-fed3-4616-a9f6-48b2c7bd0f2a
Lajoie, Patrick
29f12218-74e4-4b9f-840d-b8a085fda81c
Proud, Christopher G.
59dabfc8-4b44-4be8-a17f-578a58550cb3
Nabi, Ivan R.
cd3e0c3f-5fe9-4da2-810f-a00f3a339e7c
Roberge, Michel
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Balgi, Aruna D., Fonseca, Bruno D., Donohue, Elizabeth, Tsang, Trevor C. F., Lajoie, Patrick, Proud, Christopher G., Nabi, Ivan R. and Roberge, Michel (2009) Screen for chemical modulators of autophagy reveals novel therapeutic inhibitors of mTORC1 signaling. PLoS ONE, 4 (9), e7124.

Record type: Article

Abstract

Background
Mammalian target of rapamycin complex 1 (mTORC1) is a protein kinase that relays nutrient availability signals to control numerous cellular functions including autophagy, a process of cellular self-eating activated by nutrient depletion. Addressing the therapeutic potential of modulating mTORC1 signaling and autophagy in human disease requires active chemicals with pharmacologically desirable properties.

Methodology/Principal Findings
Using an automated cell-based assay, we screened a collection of >3,500 chemicals and identified three approved drugs (perhexiline, niclosamide, amiodarone) and one pharmacological reagent (rottlerin) capable of rapidly increasing autophagosome content. Biochemical assays showed that the four compounds stimulate autophagy and inhibit mTORC1 signaling in cells maintained in nutrient-rich conditions. The compounds did not inhibit mTORC2, which also contains mTOR as a catalytic subunit, suggesting that they do not inhibit mTOR catalytic activity but rather inhibit signaling to mTORC1. mTORC1 inhibition and autophagosome accumulation induced by perhexiline, niclosamide or rottlerin were rapidly reversed upon drug withdrawal whereas amiodarone inhibited mTORC1 essentially irreversibly. TSC2, a negative regulator of mTORC1, was required for inhibition of mTORC1 signaling by rottlerin but not for mTORC1 inhibition by perhexiline, niclosamide and amiodarone. Transient exposure of immortalized mouse embryo fibroblasts to these drugs was not toxic in nutrient-rich conditions but led to rapid cell death by apoptosis in starvation conditions, by a mechanism determined in large part by the tuberous sclerosis complex protein TSC2, an upstream regulator of mTORC1. By contrast, transient exposure to the mTORC1 inhibitor rapamycin caused essentially irreversible mTORC1 inhibition, sustained inhibition of cell growth and no selective cell killing in starvation.

Conclusion/Significance
The observation that drugs already approved for human use can reversibly inhibit mTORC1 and stimulate autophagy should greatly facilitate the preclinical and clinical testing of mTORC1 inhibition for indications such as tuberous sclerosis, diabetes, cardiovascular disease and cancer.

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Published date: 2009

Identifiers

Local EPrints ID: 142715
URI: https://eprints.soton.ac.uk/id/eprint/142715
ISSN: 1932-6203
PURE UUID: 1c4f79b5-5daa-4823-be36-29903988d44f

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Date deposited: 01 Apr 2010 09:03
Last modified: 08 Nov 2017 13:24

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Contributors

Author: Aruna D. Balgi
Author: Bruno D. Fonseca
Author: Elizabeth Donohue
Author: Trevor C. F. Tsang
Author: Patrick Lajoie
Author: Christopher G. Proud
Author: Ivan R. Nabi
Author: Michel Roberge

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