Background Colorectal cancer (CRC) is one of the most common malignancies worldwide. The intestinal microbiota plays a crucial role in regulating the tumor microenvironment (TME) of CRC. Recent studies have identified various microorganisms that inhibit tumorigenesis and progression; however, the intricate mechanisms underlying the interaction between gut microbiota and the TME require further elucidation. Previous research has shown that Lactobacillus intestinalis (L. intestinalis) can inhibit colitis, but its role in CRC remains unclear.

Methods This study aimed to evaluate the anti-tumor effects of L. intestinalis using an AOM/DSS-induced CRC model and the Apc ^min/+ spontaneous colorectal adenoma model. Immune infiltration profiles were characterized using multi-color flow cytometry. The involvement of the NOD1/NF-κB pathway was confirmed using the NOD1 receptor-specific inhibitor ML-130 and the NF-κB signaling pathway inhibitor BAY11-7082. Fecal samples and tumor tissues from patients were collected for genome DNA and mRNA profiling.

Results The abundance of L. intestinalis was decreased in fecal samples from CRC and CRA patients. Supplementation with L. intestinalis inhibited tumorigenesis in both murine models and increased the infiltration of total immune cells, particularly dendritic cells (DCs), in the TME. Co-culture of MC38 cells with L. intestinalis increased the migration ability of DCs. Mechanistically, L. intestinalis upregulated the expression of Ccl5 in MC38 cells. Knockdown of Ccl5 reduced the capacity of MC38 cells to attract DC migration when co-cultured with L. intestinalis. Moreover, ML-130 and BAY11-7082 significantly attenuated the L. intestinalis-induced CCL5 secretion, indicating the involvement of the NOD1/NF-κB pathway. In CRC patient tumor sites, the abundance of L. intestinalis positively correlated with the expression levels of CCL5, ITGAX, and NOD1 genes.

Conclusions This study provides the first evidence of the in vivo anti-tumor activity of L. intestinalis, revealing its mechanism of suppressing tumorigenesis by stimulating tumor cells to secrete CCL5 and recruiting DCs into the TME via the NOD1/NF-κB pathway. These findings highlight the significance of tumor cell responses to microbial signals within the tumor immune microenvironment, offering broader insights into the microbe-tumor-TME interaction.