Published online Oct 24, 2026. doi: 10.5306/wjco.125453
Revised: August 26, 2026
Accepted: October 8, 2026
Published online: October 24, 2026
Processing time: 107 Days and 10.1 Hours
Helicobacter pylori (H. pylori) is an important risk factor for gastric adenocarcinoma; however, clinical outcomes differ substantially according to the virulence profile of the infecting strain. Diagnostic approaches currently used generally determine pathogen status in a binary manner and therefore do not capture complex mole
To determine whether H. pylori virulence gene expression varies across clinical stages of gastric cancer (GC) and whether such variation can be used to establish a transcriptional signature associated with disease stage.
A two-tier molecular framework was applied to 400 participants recruited in Pakistan, comprising 200 GC cases and 200 non-malignant controls. Tier one examined the prevalence and risk associations of four virulence genes: Cytotoxin-associated gene A (cagA), vacuolating cytotoxin A (vacA), blood group antigen-binding adhesin A (babA), and induced by contact with epithelium gene A (iceA). Tier two quantified transcript expression in 159 H. pylori-positive GC cases spanning tumor-node-metastasis stages I-IV. Relationships between expression and clinical stage were analyzed by ordinal logistic regression, principal component analysis, and a recursive partitioning (decision tree) classifier. Receiver operating characteristic analysis and cross-validation were used to evaluate model performance.
Tier one demonstrated a distinct hierarchy among virulence factors: vacA had the strongest association [odds ratio (OR) = 22.9], with cagA ranking second (OR = 18.7). Tier two identified stage-related differences in virulence gene expression, most notably a decline in babA expression, while cagA and vacA remained comparatively elevated at later disease stages. In multivariate analysis, babA downregulation (OR = 0.536, P < 0.001) and iceA expression (OR = 0.802, P = 0.010) emerged as the most significant independent predictors of clinical stage. In internal validation, the combined four-gene signature yielded an area under the curve (AUC) of 0.769 (95%CI: 0.687-0.851) for overall stage discrimination. This apparent AUC fell to a pooled 5-fold cross-validated AUC of 0.747 (95%CI: 0.664-0.823) on held-out folds, indicating good generalizability. The decision-tree classifier separated early stages (I-II) from advanced stages (III-IV) with an apparent AUC of 0.867 (95%CI: 0.802-0.932; sensitivity 74.5%, 95%CI: 61.6-84.2%; specificity 81.7%, 95%CI: 73.2-88.0%) on the full cohort, which fell to a nested cross-validated AUC of 0.68 (95%CI: 0.59-0.77; sensitivity 54.5%, 95%CI: 41.5-67.0%; specificity 75.0%, 95%CI: 65.9-82.3%) on held-out folds.
Expression of H. pylori virulence genes differs across stages of GC. This hypothesis-generating transcriptional signature demonstrated superior discriminatory performance compared with individual gene markers in internal validation. Pending external validation in independent cohorts, it may provide molecular information that complements conventional histopathological evaluation in GC.
Core Tip: Standard diagnostic approaches for Helicobacter pylori generally establish whether the organism is present but do not resolve heterogeneity within gastric cancer. This study evaluates an expression-based discrimination model using a combined four-gene virulence transcript signature (cytotoxin-associated gene A, vacuolating cytotoxin A, blood group antigen-binding adhesin A, and induced by contact with epithelium gene A). The model identifies a stage-associated “adherence-to-invasion” transcriptional shift and, through a decision-tree classifier, discriminates early from advanced clinical stages with an area under the curve of 0.867, falling to 0.68 on nested cross-validation, a promising but modest signal warranting external validation.