Site-dependent Treg cell transcriptional reprogramming in a metastatic colorectal cancer model holds prognostic significance

Abstract In colorectal cancer (CRC), tumor-infiltrating regulatory T (Treg) cells suppress anti-tumor immunity, promoting immune evasion and tumor progression. Effective therapies require selectively targeting tumor-infiltrating Treg (TI-Treg) cells while preserving systemic Treg cells, necessitating insight into their adaptations within the tumor microenvironment. Here, CRC-organoids were implanted in the liver of Foxp3eGFP mice to investigate location-specific phenotypic differences in TI-Treg cells. Tumor tissue exhibited an increased proportion of Treg cells and a decrease of effector CD4⁺ and CD8⁺ T cells compared to matched healthy tissue. RNA sequencing of Treg cells isolated from the spleen, primary liver tumor transplant, or metastases identified gene expression profiles previously associated with CRC-related Treg cells in patients. Location-specific differences included elevated expression of WNT-pathway genes in peritoneal TI-Treg cells compared to liver counterparts. Higher expression of genes upregulated in liver TI-Treg cells correlated with poor CRC prognosis. Splenic Treg cells from tumor-bearing mice displayed distinct transcriptional profiles from both their healthy counterparts and TI-Treg cells, suggesting they represent a distinct CD4 + population. Taken together, these findings highlight TI-Treg cells heterogeneity across different tumor sites and the distinct nature of splenic Treg cells in tumor-bearing hosts.