Systematic profiling of human gut bacteria with cyclic di-AMP secretion to enhance anti-tumor immunity.

[INTRODUCTION] Gut microbiota-derived metabolites play pivotal roles in clinical tumor treatment and progression. Cyclic di-AMP serves as both a bacterial signaling molecule and an immune activator of the STING pathway. However, knowledge on cyclic di-AMP production by gut commensals remains limited, hindering the rational application of gut bacteria in cancer therapy.

[OBJECTIVES] This study systematically characterizes the metabolic profiles and genotypes associated with cyclic di-AMP synthesis in human gut commensals. We further validate the immune-activating and anti-tumor effects of high cyclic di-AMP-producing probiotics in both in vitro and in vivo non-small cell lung cancer models.

[METHODS] Cyclic di-AMP levels (intracellular and extracellular) were quantified via LC-MS in 51 representative gut bacterial species derived from 442 strains isolated from 119 human fecal samples. STING pathway activation was assessed by co-culturing THP-1 cells with supernatants from high cyclic di-AMP-producing gut probiotics. Anti-tumor efficacy was evaluated in a non-small cell lung cancer mouse model.

[RESULTS] Screening of 51 gut bacterial species identified 24 high cyclic di-AMP producers, with 18 exhibiting robust secretion capacity. Bioinformatic annotation revealed genes governing cyclic di-AMP synthesis, degradation, and secretion. Two food-grade probiotics, Limosilactobacillus fermentum DA785 and Lacticaseibacillus rhamnosus R7970, demonstrated efficient cyclic di-AMP secretion. Their supernatants significantly upregulated STING pathway-related gene expression and IFN-β secretion in THP-1 cells. Oral administration of these strains suppressed tumor growth in mice by activating immune responses within the tumor microenvironment. And Limosilactobacillus fermentum DA785 suppresses tumor growth via the STING pathway.

[CONCLUSION] This study highlights the therapeutic potential of food-grade probiotics with high cyclic di-AMP production to augment anti-tumor immunity, offering a novel microbiome-based strategy for cancer treatment.