Guo GH, Xie YB, Jiang T, An Y. Droplet digital polymerase chain reaction assay for methylated ring finger protein 180 in gastric cancer. World J Gastrointest Oncol 2022; 14(10): 2038-2047 [PMID: 36310700 DOI: 10.4251/wjgo.v14.i10.2038]
Corresponding Author of This Article
Yang An, MD, Doctor, Faculty of Hepato-Pancreato-Biliary Surgery, The Sixth Medical Center of Chinese PLA General Hospital, No. 6 Fucheng Road, Haidian District, Beijing 100048, China. dr.anyang@163.com
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Gastroenterology & Hepatology
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Observational Study
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Oct 15, 2022 (publication date) through Mar 1, 2026
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World Journal of Gastrointestinal Oncology
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1948-5204
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Guo GH, Xie YB, Jiang T, An Y. Droplet digital polymerase chain reaction assay for methylated ring finger protein 180 in gastric cancer. World J Gastrointest Oncol 2022; 14(10): 2038-2047 [PMID: 36310700 DOI: 10.4251/wjgo.v14.i10.2038]
World J Gastrointest Oncol. Oct 15, 2022; 14(10): 2038-2047 Published online Oct 15, 2022. doi: 10.4251/wjgo.v14.i10.2038
Droplet digital polymerase chain reaction assay for methylated ring finger protein 180 in gastric cancer
Guang-Hong Guo, Yi-Bin Xie, Tao Jiang, Yang An
Guang-Hong Guo, Department of Laboratory Medicine, The First Medical Center of Chinese PLA General Hospital, Beijing 100853, China
Yi-Bin Xie, Department of Pancreatic and Gastric Surgery, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, China
Tao Jiang, Medicine Innovation Research Division of Chinese PLA General Hospital, Beijing 100853, China
Yang An, Faculty of Hepato-Pancreato-Biliary Surgery, The Sixth Medical Center of Chinese PLA General Hospital, Beijing 100048, China
Author contributions: Guo GH and An Y designed the study; Guo GH and Xie YB performed the research; Guo GH and Xie YB analyzed the date; Guo GH wrote the paper; Jiang T and An Y revised the manuscript for final submission; Guo GH and Xie YB contributed equally to this study; Jiang T and An Y are the co-corresponding authors.
Supported bythe National Key Research and Development Program of China, No. 2020YFC2002700; the National Natural Science Foundation of China, No. 81972010; the CAMS Initiative for Innovative Medicine, No. 2016-I2M-1-007; and the Science Developing Funds of Navy General Hospital, No. CXPY201810.
Institutional review board statement: The study was reviewed and approved by the Ethics Committee of National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College.
Informed consent statement: The informed consent was waived by the Ethics Committee of National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Data sharing statement: No data was declared to share.
STROBE statement: The authors have read the STROBE Statement-checklist of items, and the manuscript was prepared and revised according to the STROBE Statement-checklist of items.
Corresponding author: Yang An, MD, Doctor, Faculty of Hepato-Pancreato-Biliary Surgery, The Sixth Medical Center of Chinese PLA General Hospital, No. 6 Fucheng Road, Haidian District, Beijing 100048, China. dr.anyang@163.com
Received: July 20, 2022 Peer-review started: July 20, 2022 First decision: August 20, 2022 Revised: August 21, 2022 Accepted: September 1, 2022 Article in press: September 1, 2022 Published online: October 15, 2022 Processing time: 86 Days and 1.9 Hours
Abstract
BACKGROUND
Gastric cancer (GC) is one of the most prevalent malignant tumors that endangers human health. Early diagnosis is essential for improving the prognosis and survival rate of GC patients. Ring finger protein 180 (RNF180) is involved in the regulation of cell differentiation, proliferation, apoptosis, and tumorigenesis, and aberrant hypermethylation of CpG islands in the promoter is strongly associated with the occurrence and development of GC. Thus, methylated RNF180 can be used as a potential biomarker for GC diagnosis.
AIM
To use droplet digital polymerase chain reaction (ddPCR) to quantify the methylation level of the RN180 gene. A reproducible ddPCR assay to detect methylated RNF180 from trace DNA was designed and optimized.
METHODS
The primer and probe were designed and selected, the conversion time of bisulfite was optimized, the ddPCR system was adjusted by primer concentration, amplification temperature and amplification cycles, and the detection limit of ddPCR was determined.
RESULTS
The best conversion time for blood DNA was 2 h 10 min, and that for plasma DNA was 2 h 10 min and 2 h 30 min. The results of ddPCR were better when the amplification temperature was 56 °C and the number of amplification cycles was 50. Primer concentrations showed little effect on the assay outcome. Therefore, the primer concentration could be adjusted according to the reaction system and DNA input. The assay required at least 0.1 ng of input DNA.
CONCLUSION
In summary, a ddPCR assay was established to detect methylated RNF180, which is expected to be a new diagnostic biomarker for GC.
Core Tip: Gastric cancer (GC) is one of the most prevalent malignant tumors that endangers human health. Early diagnosis is essential for improving the prognosis and survival rate of GC patients. Ring finger protein 180 (RNF180) is a newly discovered member of the ring finger protein family, and it is an important tumor suppressor gene involved in the construction of the E3 ubiquitin protein ligase in the ubiquitin proteasome system and ubiquitin degradation. Methylated RNF180 can be used as a potential biomarker for GC diagnosis. We aimed to evaluate the droplet digital polymerase chain reaction assay for methylated RNF180 in GC.
