Chen YX, He LL, Xiang XP, Shen J, Qi HY. O6-methylguanine DNA methyltransferase is upregulated in malignant transformation of gastric epithelial cells via its gene promoter DNA hypomethylation. World J Gastrointest Oncol 2022; 14(3): 664-677 [PMID: 35321285 DOI: 10.4251/wjgo.v14.i3.664]
Corresponding Author of This Article
Hong-Yan Qi, PhD, Associate Professor, Department of Pathology and Pathophysiology and Department of Radiation Oncology of the Second Affiliated Hospital, University School of Medicine, Zhejiang University, No. 866 Yuhangtang Road, Hangzhou 310058, Zhejiang Province, China. qihongyan@zju.edu.cn
Research Domain of This Article
Oncology
Article-Type of This Article
Basic Study
Open-Access Policy of This Article
This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
World J Gastrointest Oncol. Mar 15, 2022; 14(3): 664-677 Published online Mar 15, 2022. doi: 10.4251/wjgo.v14.i3.664
O6-methylguanine DNA methyltransferase is upregulated in malignant transformation of gastric epithelial cells via its gene promoter DNA hypomethylation
Yue-Xia Chen, Lu-Lu He, Hong-Yan Qi, Department of Pathology and Pathophysiology and Department of Radiation Oncology of the Second Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou 310058, Zhejiang Province, China
Yue-Xia Chen, Department of Pathology, Third Hospital of Nanchang, Nanchang 330000, Jiangxi Province, China
Xue-Ping Xiang, Department of Pathology, the Second Affiliated Hospital, Zhejiang University, Hangzhou 310058, Zhejiang Province, China
Jing Shen, Department of Pathology and Pathophysiology andDepartment of Medical Oncology of the Second Affiliated Hospital, Zhejiang University School of Medicine, Zhejiang University, Hangzhou 310058, Zhejiang Province, China
Author contributions: Chen YX and Lulu He contributed to the acquisition and analyses of the data; Xiang XP contributed to the collection and immunostaining of clinical samples; Shen J and Qi HY designed the study and made the critical revisions of the manuscript.
Supported byNational Natural Science Foundation of China, No. 81472543 and No. 81772919; Zhejiang Provincial Natural Science Foundation of China, No. LY18H160024 and No. LY20H160040.
Institutional review board statement: The study was approved by the ethics committee of Zhejiang University School of Medicine (No. 2017026).
Institutional animal care and use committee statement: The work described herein was approved by the Laboratory Animals Welfare Ethics Review Committee of Zhejiang University (No. ZJU20170522).
Conflict-of-interest statement: The authors declare no conflicts of interest for this article.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: https://creativecommons.org/Licenses/by-nc/4.0/
Corresponding author: Hong-Yan Qi, PhD, Associate Professor, Department of Pathology and Pathophysiology and Department of Radiation Oncology of the Second Affiliated Hospital, University School of Medicine, Zhejiang University, No. 866 Yuhangtang Road, Hangzhou 310058, Zhejiang Province, China. qihongyan@zju.edu.cn
Received: July 27, 2021 Peer-review started: July 27, 2021 First decision: October 3, 2021 Revised: November 10, 2021 Accepted: February 10, 2022 Article in press: February 10, 2022 Published online: March 15, 2022 Processing time: 225 Days and 21.4 Hours
Abstract
BACKGROUND
O6-methylguanine-DNA methyltransferase (MGMT) is a suicide enzyme that repairs the mispairing base O6-methyl-guanine induced by environmental and experimental carcinogens. It can transfer the alkyl group to a cysteine residue in its active site and became inactive. The chemical carcinogen N-nitroso compounds (NOCs) can directly bind to the DNA and induce the O6-methylguanine adducts, which is an important cause of gene mutation and tumorigenesis. However, the underlying regulatory mechanism of MGMT involved in NOCs-induced tumorigenesis, especially in the initiation phase, remains largely unclear.
AIM
To investigate the molecular regulatory mechanism of MGMT in NOCs-induced gastric cell malignant transformation and tumorigenesis.
METHODS
We established a gastric epithelial cell malignant transformation model induced by N-methyl-N’-nitro-N-nitrosoguanidine (MNNG) or N-methyl-N-nitroso-urea (MNU) treatment. Cell proliferation, colony formation, soft agar, cell migration, and xenograft assays were used to verify the malignant phenotype. By using quantitative real-time polymerase chain reaction (qPCR) and Western blot analysis, we detected the MGMT expression in malignant transformed cells. We also confirmed the MGMT expression in early stage gastric tumor tissues by qPCR and immunohistochemistry. MGMT gene promoter DNA methylation level was analyzed by methylation-specific PCR and bisulfite sequencing PCR. The role of MGMT in cell malignant transformation was analyzed by colony formation and soft agar assays.
RESULTS
We observed a constant increase in MGMT mRNA and protein expression in gastric epithelial cell malignant transformation induced by MNNG or MNU treatment. Moreover, we found a reduction of MGMT gene promoter methylation level by methylation-specific PCR and bisulfite sequencing PCR in MNNG/MNU-treated cells. Inhibition of the MGMT expression by O6-benzylguanine promoted the MNNG/MNU-induced malignant phenotypes. Overexpression of MGMT partially reversed the cell malignant transformation process induced by MNNG/MNU. Clinical gastric tissue analysis showed that MGMT was upregulated in the precancerous lesions and metaplasia tissues, but downregulated in the gastric cancer tissues.
CONCLUSION
Our finding indicated that MGMT upregulation is induced via its DNA promoter hypomethylation. The highly expressed MGMT prevents the NOCs-induced cell malignant transformation and tumorigenesis, which suggests a potential novel approach for chemical carcinogenesis intervention by regulating aberrant epigenetic mechanisms.
Core Tip: This study revealed a molecular regulatory mechanism of O6-methylguanine-DNA methyltransferase (MGMT) gene upregulation in the early stage of tumor development, and improved the understanding of the dynamic change of MGMT expression in different stages of tumor development, providing a new entry point for further study of the expression mechanism of key regulatory genes in the process of chemical carcinogenesis.
