PMID- 37728978 OWN - NLM STAT- PubMed-not-MEDLINE LR - 20231007 IS - 1929-0748 (Print) IS - 1929-0748 (Electronic) IS - 1929-0748 (Linking) VI - 12 DP - 2023 Sep 20 TI - Multidimensional Analysis of a Cell-Free DNA Whole Methylome Sequencing Assay for Early Detection of Gastric Cancer: Protocol for an Observational Case-Control Study. PG - e48247 LID - 10.2196/48247 [doi] LID - e48247 AB - BACKGROUND: Commonly used noninvasive serological indicators serve as a step before endoscope diagnosis and help identify the high-risk gastric cancer (GC) population. However, they are associated with high false positives and high false negatives. Alternative noninvasive approaches, such as cancer-related features in cell-free DNA (cfDNA) fragments, have been gradually identified and play essential roles in early cancer detection. The integrated analysis of multiple cfDNA features has enhanced detection sensitivity compared to individual features. OBJECTIVE: This study aimed to develop and validate an assay based on assessing genomic-scale methylation and fragmentation profiles of plasma cfDNA for early cancer detection, thereby facilitating the early diagnosis of GC. The primary objective is to evaluate the overall specificity and sensitivity of the assay in predicting GC within the entire cohort, and subsequently within each clinical stage of GC. The secondary objective involved investigating the specificity and sensitivity of the assay in combination with possible serological indicators. METHODS: This is an observational case-control study. Blood samples will be prospectively collected before gastroscopy from 180 patients with GC and 180 nonmalignant control subjects (healthy or with benign gastric diseases). Cases and controls will be randomly divided into a training and a testing data set at a ratio of 2:1. Plasma cfDNA will be isolated and extracted, followed by bisulfite-free low-depth whole methylome sequencing. A multidimensional model named Thorough Epigenetic Marker Integration Solution (THEMIS) will be constructed in the training data set. The model includes features such as the methylated fragment ratio, chromosomal aneuploidy of featured fragments, fragment size index, and fragment end motif. The performance of the model in distinguishing between patients with cancer and noncancer controls will then be evaluated in the testing data set. Furthermore, GC-related biomarkers, such as pepsinogen, gastrin-17, and Helicobacter pylori, will be measured for each patient, and their predictive accuracy will be assessed both independently and in combination with the THEMIS model. RESULTS: Recruitment began in November 2022 and will be ended in April 2024. As of August 2022,250 patients have been enrolled. The final data analysis is anticipated to be completed by September 2024. CONCLUSIONS: This is the first registered case-control study designed to investigate a stacked ensemble model integrating several cfDNA features generated from a bisulfite-free whole methylome sequencing assay. These features include methylation patterns, fragmentation profiles, and chromosomal copy number changes, with the aim of identifying the GC population. This study will determine whether multidimensional analysis of cfDNA will prove to be an effective strategy for distinguishing patients with GC from nonmalignant individuals within the Chinese population. We anticipate the THEMIS model will complement the standard-of-care screening and aid in identifying high-risk patients for further diagnosis. TRIAL REGISTRATION: ClinicalTrial.gov NCT05668910; https://www.clinicaltrials.gov/study/NCT05668910. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/48247. CI - (c)Yongjun Han, Jiangpeng Wei, Weidong Wang, Ruiqi Gao, Ning Shen, Xiaofeng Song, Yang Ni, Yulong Li, Li-Di Xu, Weizhi Chen, Xiaohua Li. Originally published in JMIR Research Protocols (https://www.researchprotocols.org), 20.09.2023. FAU - Han, Yongjun AU - Han Y AUID- ORCID: 0009-0007-3856-1144 AD - Department of General Surgery, First Hospital of Yulin, Yulin, China. FAU - Wei, Jiangpeng AU - Wei J AUID- ORCID: 0000-0001-6619-8695 AD - Department of Gastrointestinal Surgery, Xijing Hospital, Air Force Military Medical University, Xi'an, China. FAU - Wang, Weidong AU - Wang W AUID- ORCID: 0000-0003-0481-0142 AD - Department of Gastrointestinal Surgery, Xijing Hospital, Air Force Military Medical University, Xi'an, China. FAU - Gao, Ruiqi AU - Gao R AUID- ORCID: 0000-0003-0680-4256 AD - Department of Gastrointestinal Surgery, Xijing Hospital, Air Force Military Medical University, Xi'an, China. FAU - Shen, Ning AU - Shen N AUID- ORCID: 0009-0000-4690-0563 AD - Genecast Biotechnology Co, Ltd, Wuxi, China. FAU - Song, Xiaofeng AU - Song X AUID- ORCID: 0009-0009-9777-585X AD - Genecast Biotechnology Co, Ltd, Wuxi, China. FAU - Ni, Yang AU - Ni Y AUID- ORCID: 0009-0009-5180-0650 AD - Genecast Biotechnology Co, Ltd, Wuxi, China. FAU - Li, Yulong AU - Li Y AUID- ORCID: 0000-0003-0819-6987 AD - Genecast Biotechnology Co, Ltd, Wuxi, China. FAU - Xu, Li-Di AU - Xu LD AUID- ORCID: 0000-0002-3780-4972 AD - Genecast Biotechnology Co, Ltd, Wuxi, China. FAU - Chen, Weizhi AU - Chen W AUID- ORCID: 0000-0002-5099-2680 AD - Genecast Biotechnology Co, Ltd, Wuxi, China. FAU - Li, Xiaohua AU - Li X AUID- ORCID: 0000-0001-8294-7772 AD - Department of Gastrointestinal Surgery, Xijing Hospital, Air Force Military Medical University, Xi'an, China. LA - eng SI - ClinicalTrials.gov/NCT05668910 PT - Journal Article DEP - 20230920 PL - Canada TA - JMIR Res Protoc JT - JMIR research protocols JID - 101599504 PMC - PMC10551793 OTO - NOTNLM OT - chromosomal instability OT - circulating cell-free DNA OT - early detection OT - fragmentation OT - gastric cancer OT - methylation OT - multidimensional model OT - whole methylome sequencing COIS- Conflicts of Interest: NS, XS, YN, YL, LDX, and WC are employed by Genecast Biotechnology Co, Ltd. Other authors have declared no conflicts of interest. EDAT- 2023/09/20 18:42 MHDA- 2023/09/20 18:43 PMCR- 2023/09/20 CRDT- 2023/09/20 12:17 PHST- 2023/04/17 00:00 [received] PHST- 2023/08/18 00:00 [accepted] PHST- 2023/08/17 00:00 [revised] PHST- 2023/09/20 18:43 [medline] PHST- 2023/09/20 18:42 [pubmed] PHST- 2023/09/20 12:17 [entrez] PHST- 2023/09/20 00:00 [pmc-release] AID - v12i1e48247 [pii] AID - 10.2196/48247 [doi] PST - epublish SO - JMIR Res Protoc. 2023 Sep 20;12:e48247. doi: 10.2196/48247.