CDKN2A Determines Mesothelioma Cell Fate to EZH2 Inhibition

Giulia Pinton; Zhuo Wang; Cecilia Balzano; Sara Missaglia; Daniela Tavian; Renzo Boldorini; Dean A. Fennell; Martin Griffin; Laura Moro

doi:10.3389/fonc.2021.678447

CDKN2A Determines Mesothelioma Cell Fate to EZH2 Inhibition

Giulia Pinton^*, Zhuo Wang, Cecilia Balzano, Sara Missaglia, Daniela Tavian, Renzo Boldorini, Dean A. Fennell, Martin Griffin, Laura Moro^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

Abstract

Malignant pleural mesothelioma is an aggressive cancer, heterogeneous in its presentation and behaviour. Despite an increasing knowledge about molecular markers and their diagnostic and prognostic value, they are not used as much as they might be for treatment allocation. It has been recently reported that mesothelioma cells that lack BAP1 (BRCA1 Associated Protein) are sensitive to inhibition of the EZH2 (Enhancer of Zeste Homolog 2) histone methyltransferase. Since we observed strong H3K27me3 (histone H3 lysine 27 trimetylation) immunoreactivity in BAP1 wild-type mesothelioma biopsies, we decided to characterize in vitro the response/resistance of BAP1 wild-type mesothelioma cells to the EZH2 selective inhibitor, EPZ-6438. Here we demonstrate that BAP1 wild-type mesothelioma cells were rendered sensitive to EPZ-6438 upon SIRT1 (Sirtuin 1) silencing/inhibition or when cultured as multicellular spheroids, in which SIRT1 expression was lower compared to cells grown in monolayers. Notably, treatment of spheroids with EPZ-6438 abolished H3K27me3 and induced the expression of CDKN2A (Cyclin-Dependent Kinase Inhibitor 2A), causing cell growth arrest. EPZ-6438 treatment also resulted in a rapid and sustained induction of the genes encoding HIF2α (Hypoxia Inducible Factor 2α), TG2 (Transglutaminase 2) and IL-6 (Interleukin 6). Loss of CDKN2 is a common event in mesothelioma. CDKN2A silencing in combination with EPZ-6438 treatment induced apoptotic death in mesothelioma spheroids. In a CDKN2A wild-type setting apoptosis was induced by combining EPZ-6438 with 1-155, a TG2 selective and irreversible inhibitor. In conclusion, our data suggests that the expression of CDKN2A predicts cell fate in response to EZH2 inhibition and could potentially stratify tumors likely to undergo apoptosis.

Original language	English
Article number	678447
Journal	Frontiers in Oncology
Volume	11
DOIs	https://doi.org/10.3389/fonc.2021.678447
Publication status	Published - 1 Jul 2021

Bibliographical note

Copyright © 2021 Pinton, Wang, Balzano, Missaglia, Tavian, Boldorini, Fennell, Griffin and Moro. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

Funding: The authors acknowledge the financial support of project
HERMES HEreditary Risk in MESothelioma” founded by the
offer of compensation to the inhabitants of Casale Monferrato
deceased or affected by mesothelioma and Università del
Piemonte Orientale (Bando ricerca locale 2016).

Keywords

Oncology
malignant pleural mesothelioma
EZH2 inhibitor
CDKN2A/p16ink4a
TG2
multicellular spheroids

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10.3389/fonc.2021.678447Licence: CC BY 3.0

CDKN2A Determines Mesothelioma
Copyright © 2021 Pinton, Wang, Balzano, Missaglia, Tavian, Boldorini, Fennell, Griffin and Moro. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
Final published version, 5.43 MBLicence: CC BY 3.0

Cite this

@article{32664a3374a14e6bb1af7dc84a0887cc,

title = "CDKN2A Determines Mesothelioma Cell Fate to EZH2 Inhibition",

abstract = "Malignant pleural mesothelioma is an aggressive cancer, heterogeneous in its presentation and behaviour. Despite an increasing knowledge about molecular markers and their diagnostic and prognostic value, they are not used as much as they might be for treatment allocation. It has been recently reported that mesothelioma cells that lack BAP1 (BRCA1 Associated Protein) are sensitive to inhibition of the EZH2 (Enhancer of Zeste Homolog 2) histone methyltransferase. Since we observed strong H3K27me3 (histone H3 lysine 27 trimetylation) immunoreactivity in BAP1 wild-type mesothelioma biopsies, we decided to characterize in vitro the response/resistance of BAP1 wild-type mesothelioma cells to the EZH2 selective inhibitor, EPZ-6438. Here we demonstrate that BAP1 wild-type mesothelioma cells were rendered sensitive to EPZ-6438 upon SIRT1 (Sirtuin 1) silencing/inhibition or when cultured as multicellular spheroids, in which SIRT1 expression was lower compared to cells grown in monolayers. Notably, treatment of spheroids with EPZ-6438 abolished H3K27me3 and induced the expression of CDKN2A (Cyclin-Dependent Kinase Inhibitor 2A), causing cell growth arrest. EPZ-6438 treatment also resulted in a rapid and sustained induction of the genes encoding HIF2α (Hypoxia Inducible Factor 2α), TG2 (Transglutaminase 2) and IL-6 (Interleukin 6). Loss of CDKN2 is a common event in mesothelioma. CDKN2A silencing in combination with EPZ-6438 treatment induced apoptotic death in mesothelioma spheroids. In a CDKN2A wild-type setting apoptosis was induced by combining EPZ-6438 with 1-155, a TG2 selective and irreversible inhibitor. In conclusion, our data suggests that the expression of CDKN2A predicts cell fate in response to EZH2 inhibition and could potentially stratify tumors likely to undergo apoptosis.",

keywords = "Oncology, malignant pleural mesothelioma, EZH2 inhibitor, CDKN2A/p16ink4a, TG2, multicellular spheroids",

author = "Giulia Pinton and Zhuo Wang and Cecilia Balzano and Sara Missaglia and Daniela Tavian and Renzo Boldorini and Fennell, {Dean A.} and Martin Griffin and Laura Moro",

note = "Copyright {\textcopyright} 2021 Pinton, Wang, Balzano, Missaglia, Tavian, Boldorini, Fennell, Griffin and Moro. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Funding: The authors acknowledge the financial support of project HERMES HEreditary Risk in MESothelioma” founded by the offer of compensation to the inhabitants of Casale Monferrato deceased or affected by mesothelioma and Universit{\`a} del Piemonte Orientale (Bando ricerca locale 2016).",

year = "2021",

month = jul,

day = "1",

doi = "10.3389/fonc.2021.678447",

language = "English",

volume = "11",

}

TY - JOUR

T1 - CDKN2A Determines Mesothelioma Cell Fate to EZH2 Inhibition

AU - Pinton, Giulia

AU - Wang, Zhuo

AU - Balzano, Cecilia

AU - Missaglia, Sara

AU - Tavian, Daniela

AU - Boldorini, Renzo

AU - Fennell, Dean A.

AU - Griffin, Martin

AU - Moro, Laura

N1 - Copyright © 2021 Pinton, Wang, Balzano, Missaglia, Tavian, Boldorini, Fennell, Griffin and Moro. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Funding: The authors acknowledge the financial support of project HERMES HEreditary Risk in MESothelioma” founded by the offer of compensation to the inhabitants of Casale Monferrato deceased or affected by mesothelioma and Università del Piemonte Orientale (Bando ricerca locale 2016).

PY - 2021/7/1

Y1 - 2021/7/1

N2 - Malignant pleural mesothelioma is an aggressive cancer, heterogeneous in its presentation and behaviour. Despite an increasing knowledge about molecular markers and their diagnostic and prognostic value, they are not used as much as they might be for treatment allocation. It has been recently reported that mesothelioma cells that lack BAP1 (BRCA1 Associated Protein) are sensitive to inhibition of the EZH2 (Enhancer of Zeste Homolog 2) histone methyltransferase. Since we observed strong H3K27me3 (histone H3 lysine 27 trimetylation) immunoreactivity in BAP1 wild-type mesothelioma biopsies, we decided to characterize in vitro the response/resistance of BAP1 wild-type mesothelioma cells to the EZH2 selective inhibitor, EPZ-6438. Here we demonstrate that BAP1 wild-type mesothelioma cells were rendered sensitive to EPZ-6438 upon SIRT1 (Sirtuin 1) silencing/inhibition or when cultured as multicellular spheroids, in which SIRT1 expression was lower compared to cells grown in monolayers. Notably, treatment of spheroids with EPZ-6438 abolished H3K27me3 and induced the expression of CDKN2A (Cyclin-Dependent Kinase Inhibitor 2A), causing cell growth arrest. EPZ-6438 treatment also resulted in a rapid and sustained induction of the genes encoding HIF2α (Hypoxia Inducible Factor 2α), TG2 (Transglutaminase 2) and IL-6 (Interleukin 6). Loss of CDKN2 is a common event in mesothelioma. CDKN2A silencing in combination with EPZ-6438 treatment induced apoptotic death in mesothelioma spheroids. In a CDKN2A wild-type setting apoptosis was induced by combining EPZ-6438 with 1-155, a TG2 selective and irreversible inhibitor. In conclusion, our data suggests that the expression of CDKN2A predicts cell fate in response to EZH2 inhibition and could potentially stratify tumors likely to undergo apoptosis.

AB - Malignant pleural mesothelioma is an aggressive cancer, heterogeneous in its presentation and behaviour. Despite an increasing knowledge about molecular markers and their diagnostic and prognostic value, they are not used as much as they might be for treatment allocation. It has been recently reported that mesothelioma cells that lack BAP1 (BRCA1 Associated Protein) are sensitive to inhibition of the EZH2 (Enhancer of Zeste Homolog 2) histone methyltransferase. Since we observed strong H3K27me3 (histone H3 lysine 27 trimetylation) immunoreactivity in BAP1 wild-type mesothelioma biopsies, we decided to characterize in vitro the response/resistance of BAP1 wild-type mesothelioma cells to the EZH2 selective inhibitor, EPZ-6438. Here we demonstrate that BAP1 wild-type mesothelioma cells were rendered sensitive to EPZ-6438 upon SIRT1 (Sirtuin 1) silencing/inhibition or when cultured as multicellular spheroids, in which SIRT1 expression was lower compared to cells grown in monolayers. Notably, treatment of spheroids with EPZ-6438 abolished H3K27me3 and induced the expression of CDKN2A (Cyclin-Dependent Kinase Inhibitor 2A), causing cell growth arrest. EPZ-6438 treatment also resulted in a rapid and sustained induction of the genes encoding HIF2α (Hypoxia Inducible Factor 2α), TG2 (Transglutaminase 2) and IL-6 (Interleukin 6). Loss of CDKN2 is a common event in mesothelioma. CDKN2A silencing in combination with EPZ-6438 treatment induced apoptotic death in mesothelioma spheroids. In a CDKN2A wild-type setting apoptosis was induced by combining EPZ-6438 with 1-155, a TG2 selective and irreversible inhibitor. In conclusion, our data suggests that the expression of CDKN2A predicts cell fate in response to EZH2 inhibition and could potentially stratify tumors likely to undergo apoptosis.

KW - Oncology

KW - malignant pleural mesothelioma

KW - EZH2 inhibitor

KW - CDKN2A/p16ink4a

KW - TG2

KW - multicellular spheroids

UR - https://www.frontiersin.org/articles/10.3389/fonc.2021.678447/full

U2 - 10.3389/fonc.2021.678447

DO - 10.3389/fonc.2021.678447

M3 - Article

VL - 11

JO - Frontiers in Oncology

JF - Frontiers in Oncology

M1 - 678447

ER -

CDKN2A Determines Mesothelioma Cell Fate to EZH2 Inhibition

Abstract

Bibliographical note

Keywords

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