Targeting EZH1/2 induces cell cycle arrest and inhibits cell proliferation through reactivation of p57CDKN1C and TP53INP1 in mantle cell lymphoma

Wei Li, Chengfeng Bi, Yating Han, Tian Tian, Xianhuo Wang, Huijing Bao, Xiaoying Xu, Xuhan Zhang, Lu Liu, Weiwei Zhang, Hai Gao, Huaqing Wang, Huilai Zhang, Bin Meng, Xi Wang, Kai Fu

Research output: Contribution to journalArticle

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Abstract

Objective: To explore the effect of dysregulation of epigenetic regulator EZH1 and EZH2 on the proliferation in MCL and the underlying mechanisms. Methods: In this study, we elucidated the role of EZH1 and EZH2 overexpression by immunohistochemistry and correlated them to clinical outcome in 41 MCL patients. Quantitative real-time PCR and Western blot were applied to confirm the level of EZH1 and EZH2 in well-characterized MCL cell lines which were compared to those of naïve B cells. Then we manipulated the expression of EZH1 and EZH2 in MCL cells using CRISPR/Cas9 system to directly investigate their functional roles in MCL. We also evaluated the effect of two small molecule selective inhibitors, EPZ005687 and UNC1999, on MCL cell proliferation, cell cycle distribution and apoptosis in vitro. Finally, we performed RNA-sequencing (RNA-Seq) and Chromatin immunoprecipitation (ChIP) assay to further gain insight into the underlying molecular mechanisms. Results: We found that EZH2 protein is overexpressed in approximately half of this cohort of MCL cases. More importantly, the overexpression of EZH2 is associated with poor OS in the patients. Nevertheless, simple EZH2 depletion in vitro has little impact on the viability of MCL cells, predominantly because of the consequent up-regulation of EZH1. Consistently, UNC1999, a dual EZH1/2 inhibitor, unlike the EZH2 selective inhibitor EPZ005687, exerts a potent inhibitory effect on MCL cells. Furthermore, we discover CDKN1C and TP53INP1 as the two important cell cycle regulators, the expression of which are repressed by EZH1/2 mediated epigenetic regulation and are restored by EZH1/2 dual inhibition. Conclusions: Our study suggests that EZH2 participates in the pathogenesis of MCL which may serve as a potential biomarker for prognosis prediction. The dual inhibition of EZH1/2 is a promising therapeutic strategy for MCL.

Original languageEnglish (US)
Pages (from-to)530-541
Number of pages12
JournalCancer Biology and Medicine
Volume16
Issue number3
DOIs
StatePublished - Aug 1 2019

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Mantle-Cell Lymphoma
Cell Cycle Checkpoints
Epigenomics
Cell Cycle
Clustered Regularly Interspaced Short Palindromic Repeats
Cell Proliferation
RNA Sequence Analysis
Chromatin Immunoprecipitation
Real-Time Polymerase Chain Reaction
Cell Survival
B-Lymphocytes
Up-Regulation
Biomarkers
Western Blotting
Immunohistochemistry
RNA
Apoptosis
Cell Line
Proteins
In Vitro Techniques

Keywords

  • CRISPR/Cas9
  • EZH1
  • EZH2
  • Mantle cell lymphoma

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Targeting EZH1/2 induces cell cycle arrest and inhibits cell proliferation through reactivation of p57CDKN1C and TP53INP1 in mantle cell lymphoma. / Li, Wei; Bi, Chengfeng; Han, Yating; Tian, Tian; Wang, Xianhuo; Bao, Huijing; Xu, Xiaoying; Zhang, Xuhan; Liu, Lu; Zhang, Weiwei; Gao, Hai; Wang, Huaqing; Zhang, Huilai; Meng, Bin; Wang, Xi; Fu, Kai.

In: Cancer Biology and Medicine, Vol. 16, No. 3, 01.08.2019, p. 530-541.

Research output: Contribution to journalArticle

Li, Wei ; Bi, Chengfeng ; Han, Yating ; Tian, Tian ; Wang, Xianhuo ; Bao, Huijing ; Xu, Xiaoying ; Zhang, Xuhan ; Liu, Lu ; Zhang, Weiwei ; Gao, Hai ; Wang, Huaqing ; Zhang, Huilai ; Meng, Bin ; Wang, Xi ; Fu, Kai. / Targeting EZH1/2 induces cell cycle arrest and inhibits cell proliferation through reactivation of p57CDKN1C and TP53INP1 in mantle cell lymphoma. In: Cancer Biology and Medicine. 2019 ; Vol. 16, No. 3. pp. 530-541.
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abstract = "Objective: To explore the effect of dysregulation of epigenetic regulator EZH1 and EZH2 on the proliferation in MCL and the underlying mechanisms. Methods: In this study, we elucidated the role of EZH1 and EZH2 overexpression by immunohistochemistry and correlated them to clinical outcome in 41 MCL patients. Quantitative real-time PCR and Western blot were applied to confirm the level of EZH1 and EZH2 in well-characterized MCL cell lines which were compared to those of na{\"i}ve B cells. Then we manipulated the expression of EZH1 and EZH2 in MCL cells using CRISPR/Cas9 system to directly investigate their functional roles in MCL. We also evaluated the effect of two small molecule selective inhibitors, EPZ005687 and UNC1999, on MCL cell proliferation, cell cycle distribution and apoptosis in vitro. Finally, we performed RNA-sequencing (RNA-Seq) and Chromatin immunoprecipitation (ChIP) assay to further gain insight into the underlying molecular mechanisms. Results: We found that EZH2 protein is overexpressed in approximately half of this cohort of MCL cases. More importantly, the overexpression of EZH2 is associated with poor OS in the patients. Nevertheless, simple EZH2 depletion in vitro has little impact on the viability of MCL cells, predominantly because of the consequent up-regulation of EZH1. Consistently, UNC1999, a dual EZH1/2 inhibitor, unlike the EZH2 selective inhibitor EPZ005687, exerts a potent inhibitory effect on MCL cells. Furthermore, we discover CDKN1C and TP53INP1 as the two important cell cycle regulators, the expression of which are repressed by EZH1/2 mediated epigenetic regulation and are restored by EZH1/2 dual inhibition. Conclusions: Our study suggests that EZH2 participates in the pathogenesis of MCL which may serve as a potential biomarker for prognosis prediction. The dual inhibition of EZH1/2 is a promising therapeutic strategy for MCL.",
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T1 - Targeting EZH1/2 induces cell cycle arrest and inhibits cell proliferation through reactivation of p57CDKN1C and TP53INP1 in mantle cell lymphoma

AU - Li, Wei

AU - Bi, Chengfeng

AU - Han, Yating

AU - Tian, Tian

AU - Wang, Xianhuo

AU - Bao, Huijing

AU - Xu, Xiaoying

AU - Zhang, Xuhan

AU - Liu, Lu

AU - Zhang, Weiwei

AU - Gao, Hai

AU - Wang, Huaqing

AU - Zhang, Huilai

AU - Meng, Bin

AU - Wang, Xi

AU - Fu, Kai

PY - 2019/8/1

Y1 - 2019/8/1

N2 - Objective: To explore the effect of dysregulation of epigenetic regulator EZH1 and EZH2 on the proliferation in MCL and the underlying mechanisms. Methods: In this study, we elucidated the role of EZH1 and EZH2 overexpression by immunohistochemistry and correlated them to clinical outcome in 41 MCL patients. Quantitative real-time PCR and Western blot were applied to confirm the level of EZH1 and EZH2 in well-characterized MCL cell lines which were compared to those of naïve B cells. Then we manipulated the expression of EZH1 and EZH2 in MCL cells using CRISPR/Cas9 system to directly investigate their functional roles in MCL. We also evaluated the effect of two small molecule selective inhibitors, EPZ005687 and UNC1999, on MCL cell proliferation, cell cycle distribution and apoptosis in vitro. Finally, we performed RNA-sequencing (RNA-Seq) and Chromatin immunoprecipitation (ChIP) assay to further gain insight into the underlying molecular mechanisms. Results: We found that EZH2 protein is overexpressed in approximately half of this cohort of MCL cases. More importantly, the overexpression of EZH2 is associated with poor OS in the patients. Nevertheless, simple EZH2 depletion in vitro has little impact on the viability of MCL cells, predominantly because of the consequent up-regulation of EZH1. Consistently, UNC1999, a dual EZH1/2 inhibitor, unlike the EZH2 selective inhibitor EPZ005687, exerts a potent inhibitory effect on MCL cells. Furthermore, we discover CDKN1C and TP53INP1 as the two important cell cycle regulators, the expression of which are repressed by EZH1/2 mediated epigenetic regulation and are restored by EZH1/2 dual inhibition. Conclusions: Our study suggests that EZH2 participates in the pathogenesis of MCL which may serve as a potential biomarker for prognosis prediction. The dual inhibition of EZH1/2 is a promising therapeutic strategy for MCL.

AB - Objective: To explore the effect of dysregulation of epigenetic regulator EZH1 and EZH2 on the proliferation in MCL and the underlying mechanisms. Methods: In this study, we elucidated the role of EZH1 and EZH2 overexpression by immunohistochemistry and correlated them to clinical outcome in 41 MCL patients. Quantitative real-time PCR and Western blot were applied to confirm the level of EZH1 and EZH2 in well-characterized MCL cell lines which were compared to those of naïve B cells. Then we manipulated the expression of EZH1 and EZH2 in MCL cells using CRISPR/Cas9 system to directly investigate their functional roles in MCL. We also evaluated the effect of two small molecule selective inhibitors, EPZ005687 and UNC1999, on MCL cell proliferation, cell cycle distribution and apoptosis in vitro. Finally, we performed RNA-sequencing (RNA-Seq) and Chromatin immunoprecipitation (ChIP) assay to further gain insight into the underlying molecular mechanisms. Results: We found that EZH2 protein is overexpressed in approximately half of this cohort of MCL cases. More importantly, the overexpression of EZH2 is associated with poor OS in the patients. Nevertheless, simple EZH2 depletion in vitro has little impact on the viability of MCL cells, predominantly because of the consequent up-regulation of EZH1. Consistently, UNC1999, a dual EZH1/2 inhibitor, unlike the EZH2 selective inhibitor EPZ005687, exerts a potent inhibitory effect on MCL cells. Furthermore, we discover CDKN1C and TP53INP1 as the two important cell cycle regulators, the expression of which are repressed by EZH1/2 mediated epigenetic regulation and are restored by EZH1/2 dual inhibition. Conclusions: Our study suggests that EZH2 participates in the pathogenesis of MCL which may serve as a potential biomarker for prognosis prediction. The dual inhibition of EZH1/2 is a promising therapeutic strategy for MCL.

KW - CRISPR/Cas9

KW - EZH1

KW - EZH2

KW - Mantle cell lymphoma

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