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PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells

BACKGROUND: The histone methyltransferase G9a has recently been identified as a potential target for epigenetic therapy of acute myeloid leukemia (AML). However, the effect of G9a inhibition on leukemia stem cells (LSCs), which are responsible for AML drug resistance and recurrence, is unclear. In t...

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Autores principales: Jang, Ji Eun, Eom, Ju-In, Jeung, Hoi-Kyung, Chung, Haerim, Kim, Yu Ri, Kim, Jin Seok, Cheong, June-Won, Min, Yoo Hong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7158163/
https://www.ncbi.nlm.nih.gov/pubmed/32293500
http://dx.doi.org/10.1186/s13046-020-01565-3
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author Jang, Ji Eun
Eom, Ju-In
Jeung, Hoi-Kyung
Chung, Haerim
Kim, Yu Ri
Kim, Jin Seok
Cheong, June-Won
Min, Yoo Hong
author_facet Jang, Ji Eun
Eom, Ju-In
Jeung, Hoi-Kyung
Chung, Haerim
Kim, Yu Ri
Kim, Jin Seok
Cheong, June-Won
Min, Yoo Hong
author_sort Jang, Ji Eun
collection PubMed
description BACKGROUND: The histone methyltransferase G9a has recently been identified as a potential target for epigenetic therapy of acute myeloid leukemia (AML). However, the effect of G9a inhibition on leukemia stem cells (LSCs), which are responsible for AML drug resistance and recurrence, is unclear. In this study, we investigated the underlying mechanisms of the LSC resistance to G9a inhibition. METHODS: We evaluated the effects of G9a inhibition on the unfolded protein response and autophagy in AML and LSC-like cell lines and in primary CD34(+)CD38(−) leukemic blasts from patients with AML and investigated the underlying mechanisms. The effects of treatment on cells were evaluated by flow cytometry, western blotting, confocal microscopy, reactive oxygen species (ROS) production assay. RESULTS: The G9a inhibitor BIX-01294 effectively induced apoptosis in AML cell lines; however, the effect was limited in KG1 LSC-like cells. BIX-01294 treatment or siRNA-mediated G9a knockdown led to the activation of the PERK/NRF2 pathway and HO-1 upregulation in KG1 cells. Phosphorylation of p38 and intracellular generation of reactive oxygen species (ROS) were suppressed. Pharmacological or siRNA-mediated inhibition of the PERK/NRF2 pathway synergistically enhanced BIX-01294-induced apoptosis, with suppressed HO-1 expression, increased p38 phosphorylation, and elevated ROS generation, indicating that activated PERK/NRF2 signaling suppressed ROS-induced apoptosis in KG1 cells. By contrast, cotreatment of normal hematopoietic stem cells with BIX-01294 and a PERK inhibitor had no significant proapoptotic effect. Additionally, G9a inhibition induced autophagy flux in KG1 cells, while autophagy inhibitors significantly increased the BIX-01294-induced apoptosis. This prosurvival autophagy was not abrogated by PERK/NRF2 inhibition. CONCLUSIONS: PERK/NRF2 signaling plays a key role in protecting LSCs against ROS-induced apoptosis, thus conferring resistance to G9a inhibitors. Treatment with PERK/NRF2 or autophagy inhibitors could overcome resistance to G9a inhibition and eliminate LSCs, suggesting the potential clinical utility of these unique targeted therapies against AML.
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spelling pubmed-71581632020-04-21 PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells Jang, Ji Eun Eom, Ju-In Jeung, Hoi-Kyung Chung, Haerim Kim, Yu Ri Kim, Jin Seok Cheong, June-Won Min, Yoo Hong J Exp Clin Cancer Res Research BACKGROUND: The histone methyltransferase G9a has recently been identified as a potential target for epigenetic therapy of acute myeloid leukemia (AML). However, the effect of G9a inhibition on leukemia stem cells (LSCs), which are responsible for AML drug resistance and recurrence, is unclear. In this study, we investigated the underlying mechanisms of the LSC resistance to G9a inhibition. METHODS: We evaluated the effects of G9a inhibition on the unfolded protein response and autophagy in AML and LSC-like cell lines and in primary CD34(+)CD38(−) leukemic blasts from patients with AML and investigated the underlying mechanisms. The effects of treatment on cells were evaluated by flow cytometry, western blotting, confocal microscopy, reactive oxygen species (ROS) production assay. RESULTS: The G9a inhibitor BIX-01294 effectively induced apoptosis in AML cell lines; however, the effect was limited in KG1 LSC-like cells. BIX-01294 treatment or siRNA-mediated G9a knockdown led to the activation of the PERK/NRF2 pathway and HO-1 upregulation in KG1 cells. Phosphorylation of p38 and intracellular generation of reactive oxygen species (ROS) were suppressed. Pharmacological or siRNA-mediated inhibition of the PERK/NRF2 pathway synergistically enhanced BIX-01294-induced apoptosis, with suppressed HO-1 expression, increased p38 phosphorylation, and elevated ROS generation, indicating that activated PERK/NRF2 signaling suppressed ROS-induced apoptosis in KG1 cells. By contrast, cotreatment of normal hematopoietic stem cells with BIX-01294 and a PERK inhibitor had no significant proapoptotic effect. Additionally, G9a inhibition induced autophagy flux in KG1 cells, while autophagy inhibitors significantly increased the BIX-01294-induced apoptosis. This prosurvival autophagy was not abrogated by PERK/NRF2 inhibition. CONCLUSIONS: PERK/NRF2 signaling plays a key role in protecting LSCs against ROS-induced apoptosis, thus conferring resistance to G9a inhibitors. Treatment with PERK/NRF2 or autophagy inhibitors could overcome resistance to G9a inhibition and eliminate LSCs, suggesting the potential clinical utility of these unique targeted therapies against AML. BioMed Central 2020-04-15 /pmc/articles/PMC7158163/ /pubmed/32293500 http://dx.doi.org/10.1186/s13046-020-01565-3 Text en © The Author(s) 2020 Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
spellingShingle Research
Jang, Ji Eun
Eom, Ju-In
Jeung, Hoi-Kyung
Chung, Haerim
Kim, Yu Ri
Kim, Jin Seok
Cheong, June-Won
Min, Yoo Hong
PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells
title PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells
title_full PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells
title_fullStr PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells
title_full_unstemmed PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells
title_short PERK/NRF2 and autophagy form a resistance mechanism against G9a inhibition in leukemia stem cells
title_sort perk/nrf2 and autophagy form a resistance mechanism against g9a inhibition in leukemia stem cells
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7158163/
https://www.ncbi.nlm.nih.gov/pubmed/32293500
http://dx.doi.org/10.1186/s13046-020-01565-3
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