H. pylori Outer Membrane Protein Antibody Response, MHC/HLA-DMA Methylation, and Gastric Cancer Risk: A Multi-Layer Evidence Integration Study Using Mendelian Randomization

Background: Helicobacter pylori (H. pylori) infection is a well-established risk factor for gastric cancer; however, evidence on the associations between antigen-specific antibody traits, host methylation, and gastric cancer risk remains limited. Methods: Using a forward two-sample Mendelian randomization (MR) design, we integrated methylation quantitative trait loci (mQTLs), external cohorts, The Cancer Genome Atlas–Stomach Adenocarcinoma (TCGA-STAD), and functional annotations through a three-stage framework (antibody trait screening, CpG prioritization, and external replication) to evaluate a two-step pathway from host antibody response to gastric cancer via DNA methylation. Results: Among six H. pylori antigens, antibody response to outer membrane protein (OMP) showed the strongest positive association with gastric cancer risk (OR = 1.186, 95% CI: 1.079–1.304, raw p = 4.08 × 10−4). CpG-level prioritization identified cg24290947 (annotated to HLA-DMA) as a methylation site linked to gastric cancer susceptibility, with an inverse MR association (beta = −0.2347, 95% CI: −0.3980 to −0.0713, raw p = 0.00487). External exploration showed that cg24290947 methylation was elevated in H. pylori-positive gastric mucosa; however, its direct tumor–normal difference in a 12-pair cohort was not significant, although regional methylation perturbations were observed. HLA-DMA and MHC-II expression were consistently higher in tumor versus normal tissues across three cohorts, whereas TCGA-STAD did not support a stable linear relationship between cg24290947 methylation and HLA-DMA/MHC-II expression; three of six composition-adjusted models showed nominal significance but were model-dependent. These signals were enriched in antigen processing and presentation pathways. Conclusions: cg24290947/HLA-DMA represents a hypothesis-generating regional methylation candidate for further investigation rather than a confirmed linear transcriptional regulator. The MR evidence is based on European-ancestry summary data; its generalizability and any clinical predictive value warrant independent evaluation.