Clinical and Translational Research
Copyright ©The Author(s) 2022. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Jun 28, 2022; 28(24): 2705-2732
Published online Jun 28, 2022. doi: 10.3748/wjg.v28.i24.2705
Novel multiplex stool-based assay for the detection of early-stage colon cancer in a Chinese population
Hui-Hong Jiang, Si-Wei Xing, Xuan Tang, Ying Chen, Kang Lin, Lu-Wei He, Mou-Bin Lin, Er-Jiang Tang
Hui-Hong Jiang, Xuan Tang, Kang Lin, Mou-Bin Lin, Department of General Surgery, Yangpu Hospital, Tongji University, Shanghai 200090, China
Hui-Hong Jiang, Ying Chen, Lu-Wei He, Mou-Bin Lin, Er-Jiang Tang, Institute of Gastrointestinal Surgery and Translational Medicine, Tongji University School of Medicine, Shanghai 200090, China
Si-Wei Xing, Department of Urology, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China
Si-Wei Xing, Ying Chen, Kang Lin, Lu-Wei He, Mou-Bin Lin, Er-Jiang Tang, Center for Clinical Research and Translational Medicine, Yangpu Hospital, Tongji University, Shanghai 200090, China
Author contributions: Jiang HH, Xing SW and Lin K performed the experiments; Tang X, Chen Y, and Lin K participated to the collection of the clinical data and samples; Jiang HH, He LW, and Tang EJ analyzed and interpreted the data; Tang EJ, Lin MB and He LW conceived and managed the study; Jiang HH and Xing SW drafted the manuscript; and all authors have read and approve the final manuscript.
Supported by Shanghai Pujiang Program, No. 21PJD066; Shanghai Municipal Commission of Health and Family Planning, No. ZK2019A19; Shanghai Municipal Science and Technology Commission, No. 19411971500; and Shanghai Yangpu District Science and Technology Commission, No. YPM202101.
Institutional review board statement: This study was reviewed and approved by the Ethics Committee of Yangpu Hospital, Tongji University (LL-2018-SCI-003).
Clinical trial registration statement: This study was registered at the Chinese Clinical Trial Registry (ChiCTR1800019552).
Informed consent statement: All study participants or their legal guardian provided informed written consent about personal and medical data collection prior to study enrolment.
Conflict-of-interest statement: The authors have no conflicts of interest to declare.
Data sharing statement: No additional data are available.
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: Er-Jiang Tang, MSc, Statistician, Center for Clinical Research and Translational Medicine, Yangpu Hospital, Tongji University, No. 450 Tengyue Road, Shanghai 200090, China. tangerjiang1988051@163.com
Received: December 4, 2021
Peer-review started: December 4, 2021
First decision: January 8, 2022
Revised: January 14, 2022
Accepted: May 13, 2022
Article in press: May 13, 2022
Published online: June 28, 2022
Processing time: 202 Days and 4.6 Hours
Abstract
BACKGROUND

Stool DNA (sDNA) methylation analysis is a promising, noninvasive approach for colorectal cancer screening; however, reliable biomarkers for detecting early-stage colon cancer (ECC) are lacking, particularly in the Chinese population.

AIM

To identify a novel stool-based assay that can improve the effectiveness of ECC screening.

METHODS

A blinded case-control study was performed using archived stool samples from 125 ECC patients, and 125 control subjects with normal colonoscopy. The cohort was randomly divided into training and test sets at a 1.5:1 ratio. Targeted bisulfite sequencing (TBSeq) was conducted on five pairs of preoperative and postop-erative sDNA samples from ECC patients to identify DNA methylation biomarkers, which were validated using pyrosequencing. By logistic regression analysis, a multiplex stool-based assay was developed in the training set, and the detection performance was further assessed in the test set and combined set. The χ2 test was used to investigate the association of detection sensitivity with clinico-pathological features.

RESULTS

Following TBSeq, three hypermethylated cytosine-guanine sites were selected as biomarkers, including paired box 8, Ras-association domain family 1 and secreted frizzled-related protein 2, which differed between the groups and were involved in important cancer pathways. An sDNA panel containing the three biomarkers was constructed with a logistic model. Receiver operating characteristic (ROC) analysis revealed that this panel was superior to the fecal immunochemical test (FIT) or serum carcinoembryonic antigen for the detection of ECC. We further found that the combination of the sDNA panel with FIT could improve the screening effectiveness. In the combined set, the sensitivity, specificity and area under the ROC curve for this multiplex assay were 80.0%, 93.6% and 0.918, respectively, and the performance remained excellent in the subgroup analysis by tumor stage. In addition, the detection sensitivity did not differ with tumor site, tumor stage, histological differentiation, age or sex, but was significantly higher in T4 than in T1-3 stage tumors (P = 0.041).

CONCLUSION

We identified a novel multiplex stool-based assay combining sDNA methylation biomarkers and FIT, which could detect ECC with high sensitivity and specificity throughout the colon, showing a promising application perspective.

Keywords: Colon cancer; Early screening; Stool biomarker; DNA methylation; Fecal immunochemical test

Core Tip: Stool DNA (sDNA) methylation analysis has a promising application in the early diagnosis of colorectal cancer. However, reliable biomarkers for detecting early-stage colon cancer (ECC) are lacking. In this study, by targeted bisulfite sequencing, we identified a novel multiplex stool-based assay combining three sDNA methylation biomarkers and fecal immunochemical test. Further validation in larger samples by pyrosequencing showed that it enabled the diagnosis of ECC with high sensitivity and specificity throughout the colon. To our knowledge, this is the first study to focus on ECC screening in China.