A Multiomics Analysis of the Close Connection between Intratumoral Microbiota and Immune Cell Infiltration in Colorectal Cancer

Abstract

The colorectal intratumoral microbiome and its association with the expression of the tumor genome and immune cell infiltration remain poorly characterized. Our study aims to investigate the relationship between intratumoral microbiota with tumor immune infiltration, patient prognosis, and potential downstream signaling pathways. We collected biopsy samples of tumor tissue and paracancerous tissue from 92 patients with colorectal cancer, and acquired microbiota profiling in these samples using 16s rRNA sequencing. Meanwhile, the immune markers including CD8, FOXP3, CD163, PD-1 and PD-L1 were stained by immunohistochemistry (IHC) to identify the immune infiltration in tumors. Furthermore, we used The Cancer Genome Atlas and The Cancer Microbiome Atlas databases to conduct multiomics analysis on tumor flora and patient survival, tumor gene expression profile and potential downstream pathways. There was a significant difference in α-diversity (p = 0.00051) and β-diversity (p = 0.004) between tumor and paracancerous tissues. The β-diversity of intratumoral bacterial differed by colorectal cancer tumor stage (early vs. late stage, p = 0.049) and location (left vs. right colon, p = 0.04). Stage-related flora cluster (Porphyromonas, Lachnoclostridium, Bacteroides, Aggregatibacter, and Hungatella) were identified and found to be associated with poor prognosis in colorectal cancer patients (HR = 1.79, p = 0.015). By IHC staining, we found that expression of PD-1 and FOXP3 was significantly reduced at low abundance of stage-related bacterial cluster (p<0.05). Among of them, Hungatella was negatively associated with CD8+T cell infiltration (p<0.05) in tumor. Besides, tumor-location related flora cluster (Bacteroides and Blautia) were identified and found to be associated with good prognosis in colorectal cancer patients (HR = 0.52, p = 0.011). Expression of CD163 was decreased at high abundance of location-related bacterial cluster (p<0.05). Among of them, Blautia was negatively correlated with tumor-associated macrophage infiltration(p<0.05). Furthermore, we found that the stage-related flora cluster was positively connected with the pathway of bile acid metabolism, whereas the location-dependent cluster was negatively correlated with this pathway. We found specific intratumoral bacterial clusters that were related to tumor stage and location, and the clusters were strongly associated with tumor immune infiltration and patient prognosis. Our findings may provide new viewpoint for future research between intratumoral microbiota, metabolism pathway and tumor microenvironment.