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GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells

The process of autophagy and its role in survival of human neuroblastoma cell cultures was studied upon addition of an anti-GD2 ganglioside (GD2) 14G2a mouse monoclonal antibody (14G2a mAb) and an aurora A kinase specific inhibitor, MK-5108. It was recently shown that combination of these agents sig...

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Autores principales: Durbas, Małgorzata, Pabisz, Paweł, Wawak, Katarzyna, Wiśniewska, Aneta, Boratyn, Elżbieta, Nowak, Iwona, Horwacik, Irena, Woźnicka, Olga, Rokita, Hanna
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer US 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6153511/
https://www.ncbi.nlm.nih.gov/pubmed/30027525
http://dx.doi.org/10.1007/s10495-018-1472-9
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author Durbas, Małgorzata
Pabisz, Paweł
Wawak, Katarzyna
Wiśniewska, Aneta
Boratyn, Elżbieta
Nowak, Iwona
Horwacik, Irena
Woźnicka, Olga
Rokita, Hanna
author_facet Durbas, Małgorzata
Pabisz, Paweł
Wawak, Katarzyna
Wiśniewska, Aneta
Boratyn, Elżbieta
Nowak, Iwona
Horwacik, Irena
Woźnicka, Olga
Rokita, Hanna
author_sort Durbas, Małgorzata
collection PubMed
description The process of autophagy and its role in survival of human neuroblastoma cell cultures was studied upon addition of an anti-GD2 ganglioside (GD2) 14G2a mouse monoclonal antibody (14G2a mAb) and an aurora A kinase specific inhibitor, MK-5108. It was recently shown that combination of these agents significantly potentiates cytotoxicity against IMR-32 and CHP-134 neuroblastoma cells in vitro, as compared to the inhibitor used alone. In this study we gained mechanistic insights on autophagy in the observed cytotoxic effects exerted by both agents using cytotoxicity assays, RT-qPCR, immunoblotting, and autophagy detection methods. Enhancement of the autophagy process in the 14G2a mAb- and MK-5108-treated IMR-32 cells was documented by assessing autophagic flux. Application of a lysosomotropic agent—chloroquine (CQ) affected the 14G2a mAb- and MK-5108-stimulated autophagic flux. It is our conclusion that the 14G2a mAb (40 μg/ml) and MK-5108 inhibitor (0.1 μM) induce autophagy in IMR-32 cells. Moreover, the combinatorial treatment of IMR-32 cells with the 14G2a mAb and CQ significantly potentiates cytotoxic effect, as compared to CQ used alone. Most importantly, we showed that interfering with autophagy at its early and late step augments the 14G2a mAb-induced apoptosis, therefore we can conclude that inhibition of autophagy is the primary mechanism of the CQ-mediated sensitization to the 14G2a mAb-induced apoptosis. Although, there was no virtual stimulation of autophagy in the 14G2a mAb-treated CHP-134 neuroblastoma cells, we were able to show that PHLDA1 protein positively regulates autophagy and this process exists in a mutually exclusive manner with apoptosis in PHLDA1-silenced CHP-134 cells. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10495-018-1472-9) contains supplementary material, which is available to authorized users.
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spelling pubmed-61535112018-10-09 GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells Durbas, Małgorzata Pabisz, Paweł Wawak, Katarzyna Wiśniewska, Aneta Boratyn, Elżbieta Nowak, Iwona Horwacik, Irena Woźnicka, Olga Rokita, Hanna Apoptosis Article The process of autophagy and its role in survival of human neuroblastoma cell cultures was studied upon addition of an anti-GD2 ganglioside (GD2) 14G2a mouse monoclonal antibody (14G2a mAb) and an aurora A kinase specific inhibitor, MK-5108. It was recently shown that combination of these agents significantly potentiates cytotoxicity against IMR-32 and CHP-134 neuroblastoma cells in vitro, as compared to the inhibitor used alone. In this study we gained mechanistic insights on autophagy in the observed cytotoxic effects exerted by both agents using cytotoxicity assays, RT-qPCR, immunoblotting, and autophagy detection methods. Enhancement of the autophagy process in the 14G2a mAb- and MK-5108-treated IMR-32 cells was documented by assessing autophagic flux. Application of a lysosomotropic agent—chloroquine (CQ) affected the 14G2a mAb- and MK-5108-stimulated autophagic flux. It is our conclusion that the 14G2a mAb (40 μg/ml) and MK-5108 inhibitor (0.1 μM) induce autophagy in IMR-32 cells. Moreover, the combinatorial treatment of IMR-32 cells with the 14G2a mAb and CQ significantly potentiates cytotoxic effect, as compared to CQ used alone. Most importantly, we showed that interfering with autophagy at its early and late step augments the 14G2a mAb-induced apoptosis, therefore we can conclude that inhibition of autophagy is the primary mechanism of the CQ-mediated sensitization to the 14G2a mAb-induced apoptosis. Although, there was no virtual stimulation of autophagy in the 14G2a mAb-treated CHP-134 neuroblastoma cells, we were able to show that PHLDA1 protein positively regulates autophagy and this process exists in a mutually exclusive manner with apoptosis in PHLDA1-silenced CHP-134 cells. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s10495-018-1472-9) contains supplementary material, which is available to authorized users. Springer US 2018-07-19 2018 /pmc/articles/PMC6153511/ /pubmed/30027525 http://dx.doi.org/10.1007/s10495-018-1472-9 Text en © The Author(s) 2018 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.
spellingShingle Article
Durbas, Małgorzata
Pabisz, Paweł
Wawak, Katarzyna
Wiśniewska, Aneta
Boratyn, Elżbieta
Nowak, Iwona
Horwacik, Irena
Woźnicka, Olga
Rokita, Hanna
GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells
title GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells
title_full GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells
title_fullStr GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells
title_full_unstemmed GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells
title_short GD2 ganglioside-binding antibody 14G2a and specific aurora A kinase inhibitor MK-5108 induce autophagy in IMR-32 neuroblastoma cells
title_sort gd2 ganglioside-binding antibody 14g2a and specific aurora a kinase inhibitor mk-5108 induce autophagy in imr-32 neuroblastoma cells
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6153511/
https://www.ncbi.nlm.nih.gov/pubmed/30027525
http://dx.doi.org/10.1007/s10495-018-1472-9
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