Calcium overload via PVT1 reprograms neutrophil fate and constrains gastric cancer progression.

BACKGROUND: The aberrant upregulation of PVT1 in gastric cancer (GC) has emerged as a critical molecular marker, while its mechanism of remodeling the tumor microenvironment (TME) through metabolic regulation remains unclear. This study aims to unveil the novel mechanism by which PVT1 regulates the fate determination of tumor-associated neutrophils (TANs) via the calcium signaling pathway. METHODS: Clinical analysis of 90 GC cases revealed a correlation between PVT1 expression and neutrophil infiltration in tumor tissues. Mechanistically, multi-omics integration of transcriptomics and 4D-label-free quantitative proteomics identified PVT1-associated pathways driving GC progression. Functional validation through in vitro and in vivo models demonstrated PVT1's oncogenic role, while transcriptomic profiling further decoded neutrophil activation mechanisms regulated by PVT1. RESULTS: The expression of PVT1 was significantly elevated in GC tissues compared to adjacent non-tumor tissues (p < 0.001), with 73.3% of cases showing an increased proportion of PVT1-positive cells. High PVT1 expression negatively correlated with the infiltration of neutrophils (r = - 0.3554, p = 0.0012), and patients in the PVT1 + CD66b- subgroup exhibited the poorest prognosis, characterized by a median survival of 20.5 months and a mortality rate of 72.5%. Silencing PVT1 upregulated calcium signaling pathway-related genes (RYR2 and RYR3) in GC cells, accompanied by a marked increase in intracellular Ca(2)⁺ concentration (p < 0.01). Proteomic analysis revealed that PVT1 knockdown significantly enhanced the secretion of synaptopodin (SYNPO), which demonstrated co-expression and physical interactions with calcium channel proteins RYR2/RYR3. PVT1 knockdown markedly suppressed GC cell proliferation, migration, and invasion (p < 0.05) while elevating the expression of γ-H2AX (p < 0.05). In xenograft models, tumors with PVT1 knockdown displayed reduced volume and decreased Ki67 expression (p < 0.01). Conditioned medium from PVT1-knockdown GC cells promoted neutrophil activation, delayed apoptosis, and amplified intracellular Ca(2)⁺ signaling. Exogenous SYNPO supplementation mimicked these effects and attenuated neutrophil replicative senescence by modulating p53 pathway-associated genes (TP53, CHEK1, and SERPINE1). CONCLUSIONS: Overall, this study provides the first evidence that PVT1 establishes a metabolic checkpoint by inducing calcium overload, thereby regulating both the senescence process and functional phenotype of neutrophils, which provides a novel metabolic intervention target for GC immunotherapy.