Helicobacter Pylori Infection Status and Pathological Features for Predicting Gastric Cancer Biological Behavior and Prognosis: A Real-World Observational Study

Background and Rationale:

Helicobacter pylori (H. pylori) infection is a major risk factor for gastric cancer and is closely associated with chronic gastritis, mucosal atrophy, intestinal metaplasia, dysplasia, and gastric carcinogenesis. Although H. pylori eradication can reduce the risk of gastric cancer, some patients still develop gastric cancer after eradication therapy, and patients treated with endoscopic submucosal dissection (ESD) may remain at risk for local recurrence, metachronous gastric cancer, and other gastric neoplastic lesions during follow-up.

In clinical practice, H. pylori status can be evaluated by several methods, including the 13C-urea breath test, serum H. pylori antibody testing, and pathological assessment of gastric tissue. These methods reflect different biological and clinical information. The 13C-urea breath test mainly reflects current active infection, serum antibody testing may reflect previous or current exposure, and pathological assessment provides local tissue-based information from specific gastric sites.

However, these test results may be inconsistent. H. pylori colonization can be patchy and unevenly distributed within the stomach. In patients with early gastric cancer, local tumor-related changes, mucosal atrophy, intestinal metaplasia, spasmolytic polypeptide-expressing metaplasia, prior eradication therapy, and changes in the gastric mucosal microenvironment may influence bacterial density and detectability. Therefore, a negative pathological finding in the tumor area does not necessarily exclude previous H. pylori exposure or H. pylori-related background mucosal changes.

Current evidence suggests that H. pylori may be more frequently detected in non-tumor or peritumoral mucosa than in tumor tissue itself, and that H. pylori-related mucosal changes may be associated with specific pathological phenotypes of early gastric cancer. Nevertheless, the clinical meaning of discordant H. pylori test patterns and the relationship among H. pylori intragastric distribution, gastric pathological characteristics, post-eradication gastric cancer, and metachronous gastric cancer remain insufficiently understood.

Study Objective:

The overall objective of this study is to establish a sequential clinical-pathological framework linking Helicobacter pylori (H. pylori) test heterogeneity, pathological features including tissue-based and intragastric H. pylori distribution, gastric cancer subtype and biological behavior, and long-term prognosis in patients with early gastric cancer or related gastric neoplastic lesions undergoing endoscopic submucosal dissection (ESD).

The study will focus on four linked components:

• To characterize heterogeneity among different H. pylori testing results, including 13C-urea breath test, serum H. pylori antibody testing, and pathological assessment of gastric tissue. Concordant and discordant testing patterns will be used to define current infection, previous exposure, post-eradication status, no evidence of infection, and tissue-based H. pylori detection.
• To evaluate pathological features associated with these H. pylori testing patterns, including tissue-based and intragastric H. pylori distribution in tumor tissue, lesion-adjacent mucosa, and non-lesion background mucosa when available. Background mucosal changes, including chronic inflammation, active inflammation, atrophy, intestinal metaplasia, dysplasia, and other pathology-reported mucosal findings, will also be assessed. Lesion-level pathological features, including lesion location, histological subtype, differentiation, depth of invasion, lymphovascular invasion, and margin status, will be recorded.
• To classify and compare different gastric cancer subtypes and biological behavior patterns according to the preceding H. pylori testing and pathological features. These subtypes may include current or persistent H. pylori infection-associated gastric cancer, past exposure or post-eradication gastric cancer, gastric cancer with no evidence of H. pylori infection, proximal versus distal gastric cancer, differentiated versus undifferentiated histology, and solitary versus synchronous or multifocal early gastric cancer detected within 1 year when applicable.
• To evaluate long-term clinical outcomes after ESD, with particular attention to whether the integrated analysis of H. pylori infection status and pathological features can predict long-term outcomes in early gastric cancer, including local recurrence, metachronous gastric cancer, additional surgical treatment, survival, and other clinically meaningful outcomes.

Study Design:

This is a multicenter retrospective and prospective real-world observational cohort study led by Chinese PLA General Hospital and conducted at participating medical centers. Approximately 1500 participants will be included. About 1000 participants will be retrospectively identified from existing clinical records, endoscopic databases, H. pylori testing records, ESD pathological records, and follow-up information. About 500 additional participants are planned to be prospectively enrolled.

Eligible participants will be patients with early gastric cancer, high-grade intraepithelial neoplasia, or other gastric neoplastic lesions treated with or scheduled for ESD according to standard clinical indications.

This study is observational. Participants will not be assigned to any experimental intervention by the study protocol. All diagnostic evaluation, H. pylori testing, ESD treatment, perioperative management, pathological assessment, H. pylori eradication therapy when clinically indicated, and follow-up will be performed according to routine clinical practice and institutional standards.

H. pylori Assessment:

For retrospectively included participants, available H. pylori testing results will be extracted from existing medical records, including 13C-urea breath test, serum H. pylori antibody testing, pathological assessment, prior eradication history, eradication confirmation, and follow-up H. pylori testing results when available.

For prospectively enrolled participants, H. pylori assessment will be performed according to routine clinical practice, including serum H. pylori IgG antibody testing and the 13C-urea breath test before ESD when clinically indicated. Pathological assessment of gastric tissue will be collected from routine clinical specimens when available, including diagnostic biopsy specimens and ESD specimens.

Participants will be categorized according to H. pylori infection and eradication patterns, including H. pylori-positive gastric cancer without prior eradication, persistent H. pylori-positive gastric cancer after eradication, H. pylori-negative gastric cancer after eradication, and H. pylori-negative gastric cancer with no evidence of infection.

Intragastric Distribution and Background Mucosal Assessment:

The study will record the anatomical location of gastric lesions and available tissue-based findings by gastric region. Locations may include the cardia, fundus, gastric body, gastric angle, and antrum, as well as lesser curvature, greater curvature, anterior wall, and posterior wall when available.

When tissue material allows, pathological assessment will evaluate H. pylori detection in tumor tissue, lesion-adjacent mucosa, and non-lesion background mucosa. Background mucosal changes, including chronic inflammation, active inflammation, atrophy, intestinal metaplasia, and other pathology-reported mucosal findings, will be recorded when available. These findings will be used to explore whether discordant H. pylori test patterns can be explained by intragastric distribution, local tissue detectability, or background mucosal changes.

Endoscopic and Pathological Assessment:

Endoscopic data will include lesion location, lesion size, macroscopic morphology, surface characteristics, ulceration or scar findings, and other relevant endoscopic features.

Pathological data will include histological diagnosis, histological subtype, degree of differentiation, depth of invasion, lymphovascular invasion, horizontal and vertical margin status, and whether curative resection criteria are met when applicable. The study will evaluate whether H. pylori infection status, prior eradication status, discordant testing patterns, and intragastric distribution are associated with gastric cancer pathological features and background mucosal changes.

Perioperative Outcomes:

Although the main focus of this study is H. pylori status, intragastric distribution, pathological characteristics, and long-term prognosis, ESD-related perioperative outcomes will also be collected as clinically relevant supportive data. These may include intraoperative bleeding requiring endoscopic hemostasis, procedure duration, intraoperative perforation, en bloc resection, R0 resection, delayed bleeding, postoperative complications, and length of hospital stay.

Long-term Follow-up:

Participants will be followed after ESD according to routine clinical practice. For retrospectively included participants, available follow-up information at approximately 1, 2, and 3 years after ESD and longer-term follow-up when available will be extracted from medical records, endoscopic follow-up records, pathological reports, telephone follow-up, and survival information. For prospectively enrolled participants, follow-up will be conducted according to routine clinical practice.

Follow-up evaluations may include outpatient visits, telephone follow-up, endoscopic examination, imaging examination when clinically indicated, and review of medical records. Follow-up outcomes will be assessed at 1, 2, and 3 years after ESD and during long-term follow-up through study completion. Outcomes will include local residual disease, local recurrence, synchronous or multifocal early gastric cancer detected within 1 year, metachronous gastric cancer, additional surgical treatment, survival status, and other clinically relevant outcomes.

Follow-up endoscopy will usually be performed every 6 to 12 months according to clinical practice and individual risk. H. pylori eradication therapy, when clinically indicated, will be recorded, as will subsequent changes in H. pylori status when follow-up testing is available.

Analysis Plan:

The study will first summarize H. pylori test patterns based on 13C-urea breath test, serum H. pylori antibody testing, and pathological assessment. Concordance and discordance among these testing modalities will be evaluated using overall agreement rates, pairwise agreement rates, and agreement statistics when appropriate.

The study will then describe H. pylori intragastric and tissue-based distribution and related background mucosal changes by gastric region and pathological compartment. Associations between H. pylori test patterns, prior eradication status, tissue distribution, background mucosal changes, and gastric pathological features will be evaluated using appropriate statistical methods. Categorical variables will be compared using chi-square or Fisher's exact tests, and continuous variables will be compared using parametric or non-parametric tests depending on distribution.

For long-term outcomes, local recurrence, metachronous gastric cancer, additional surgical treatment, recurrence-free survival, overall survival, and other clinically meaningful outcomes will be assessed at 1, 2, and 3 years after ESD and during long-term follow-up through study completion. Logistic regression or Cox proportional hazards models may be used to identify factors associated with these outcomes when sufficient events are available. Potential covariates may include age, sex, study center, retrospective versus prospective data source, lesion location, lesion size, histological type, differentiation, depth of invasion, lymphovascular invasion, margin status, atrophy or intestinal metaplasia, H. pylori testing pattern, tissue-based H. pylori distribution, and prior eradication history.

Study Significance:

This study is intended to provide a systematic clinical and pathological evaluation of H. pylori infection status, discordant test patterns, tissue-based and intragastric distribution, pathological features, gastric cancer biological behavior, and long-term outcomes in patients undergoing ESD for early gastric cancer or related gastric neoplastic lesions.

The findings may help improve interpretation of H. pylori test results, clarify the pathological basis of discordant H. pylori detection, and identify H. pylori-related background mucosal changes. In addition, this study aims to characterize the pathological features and biological behavior of gastric cancer after H. pylori eradication and other H. pylori-related gastric cancer subtypes.

By integrating H. pylori infection status, prior eradication status, tissue-based H. pylori distribution, background mucosal changes, and tumor pathological characteristics, this study may support subtype-specific prognostic evaluation and risk-adapted surveillance strategies for local recurrence, metachronous gastric cancer, additional surgical treatment, and other clinically meaningful outcomes after ESD.

Inclusion criteria

• Adults aged 18 to 80 years
• Retrospectively identified or prospectively enrolled patients with early gastric cancer, high-grade intraepithelial neoplasia, or other gastric neoplastic lesions treated with or scheduled for endoscopic submucosal dissection according to standard clinical indications at participating medical centers
• Availability or planned availability of sufficient Helicobacter pylori assessment data for study classification, including 13C-urea breath test, serum Helicobacter pylori IgG antibody testing, pathological assessment of gastric tissue, prior H. pylori infection history, prior eradication history, eradication confirmation, or follow-up H. pylori testing results when available
• Availability or planned availability of gastric tissue pathological assessment from routine biopsy specimens and/or endoscopic submucosal dissection specimens for evaluation of tissue-based H. pylori detection, background mucosal changes, lesion pathological characteristics, and ESD-related pathological findings
• Availability or planned availability of key clinical, endoscopic, and pathological information required to evaluate gastric cancer subtype, biological behavior, and curability after ESD, including lesion location, histological subtype, differentiation, depth of invasion, lymphovascular invasion, margin status, curative resection status, and additional treatment information when available
• Availability of follow-up data for retrospectively included participants, or willingness and ability to participate in follow-up evaluations for prospectively enrolled participants, including endoscopic follow-up at approximately 1, 2, and 3 years after ESD and longer-term follow-up when available
• Adequate cardiac, hepatic, renal, and pulmonary function to tolerate endoscopic treatment and routine follow-up for prospectively enrolled participants scheduled for ESD
• Ability to provide written informed consent for prospectively enrolled participants, or availability of ethics-approved retrospective clinical data for retrospectively included participants

Exclusion criteria

• Patients whose final diagnosis does not meet the study population definition, including those without early gastric cancer, high-grade intraepithelial neoplasia, or other eligible gastric neoplastic lesions
• Insufficient clinical, H. pylori testing, endoscopic, pathological, or follow-up information to classify H. pylori infection or eradication status or to evaluate the main study outcomes
• Lack of adequate pathological material or pathological information for assessment of gastric lesion characteristics, tissue-based H. pylori detection, or background mucosal changes
• Previous gastrectomy or major gastric surgery that substantially alters gastric anatomy and prevents reliable assessment of lesion location, intragastric distribution, or background mucosal status
• For prospectively enrolled participants scheduled for ESD, severe comorbid conditions, such as uncontrolled cardiovascular or cerebrovascular disease, severe coagulopathy, or very poor general condition, that make endoscopic submucosal dissection unsafe
• For prospectively enrolled participants scheduled for ESD, anticoagulant or antiplatelet therapy that cannot be safely interrupted or managed during the perioperative period, or active bleeding tendency judged to significantly increase procedural risk
• Pregnancy or breastfeeding for prospectively enrolled participants
• Inability or unwillingness to comply with study procedures or follow-up requirements for prospectively enrolled participants
• Any other condition judged by the investigators to make the participant unsuitable for the study