Impact of <i>Helicobacter pylori</i> infection on molecular alterations and immune dynamics in gastric cancer.

Abstract

2626 Background: Helicobacter pylori (HP) infection, a major risk factor for gastric cancer (GC), modulates tumor immunity. Evidence indicates that H. pylori infection status correlates with the efficacy of immune checkpoint inhibitors (ICI), with varying outcomes in H. pylori -positive ( HPP ) and -negative ( HPN ) patients. This study investigated the impact of HP infection on splicing alterations to elucidate its role in shaping immune responses and the tumor immune microenvironment (TIME), immune checkpoint molecule expression, transcriptional profiles in GC. Methods: Tumors and adjacent normal tissues were collected from 24 patients with GC, comprising 39 tumor samples from HPP patients, 27 samples from HPN patients, and 13 and 10 normal samples of each, respectively. RNA sequencing was performed on these tissues and whole blood RNA from six patients (four HPP and two HPN ) to analyze the alternative splicing (AS) events, the transcriptional profiles, and immune-related gene expression. Differential gene expression (DGE) and enrichment analyses were conducted, and immune cell fractions were evaluated using CIBERSORTx. Results: DGE analysis revealed that HPP tumors were enriched in genes related to cell cycle regulation, whereas HPN tumors were enriched in immune response pathways, including those involved in leukocyte activation, chemokine signaling, and immune effector processes. Additionally, HPN tumors showed higher expression of immune checkpoint molecules, such as CD160 (p = 0.016), PDCD1LG2 (p = 0.0082), and BTLA (p = 0.025). Immune cell profiling demonstrated increased proportions of gamma-delta T cells (p = 0.0077), resting dendritic cells (p = 0.0002), and neutrophils (p = 0.016), reflecting enhanced immune activation and a favorable ICI response. In contrast, HPP tumors were enriched in cell cycle-related pathways, suggesting a proliferative phenotype. HPP tumors also exhibited higher levels of M0 macrophages (p = 0.0039) and CD276 expression (p = 0.0082), indicative of an immunosuppressive TIME. AS analysis identified increased intron retention (IR) events in HPP tumors, particularly in genes associated with RNA processing and extracellular matrix remodeling. These alterations may contribute to immune evasion and tumor progression. In the peripheral blood, HPP samples exhibited upregulation of tripartite motif family genes, which are implicated in immune modulation. Conclusions: This study demonstrated that HP infection significantly affects the TME and gene expression profiles of GC. HPP tumors are characterized by increased M0 macrophage populations, CD276 expression, and IR events that contribute to immunosuppression and tumor progression. In contrast, HPN tumors exhibit greater immune activation and checkpoint molecule diversity. These findings highlight the potential role of HP status in shaping the immune landscape of GC and influencing responsiveness to ICI.