Fibroblasts selectively modulate immune cell-derived cytokine effects on tumor cells in colorectal cancer

Abstract

Introduction Colorectal cancer (CRC) is a heterogeneous malignancy and a major cause of cancer-related mortality worldwide. Cancer-associated fibroblasts (CAFs) accumulate in tumors and correlate with poor patient survival, suggesting a central role in immune regulation. Patient-derived organoids (PDO) maintain the intra-tumoral cellular heterogeneity of the original tissue, thus, they represent one of the best methods to study human cancers. The tumor microenvironment (TME) contains diverse immune cell populations, including innate lymphoid cells (ILCs), yet how stromal components influence cytokine-driven tumor–immune interactions remains unclear. Methods PDOs from CRC patients were used to screen TME-derived cytokines affecting tumor growth. Organoid-forming efficiency and signaling pathway activation were analyzed following cytokine stimulation in the presence or absence of CAFs. Tumor-infiltrating ILC subsets were characterized in patient samples, and co-culture systems were employed to assess cytokine production and stromal–immune interactions. Results IL-22 was identified as a cytokine that increased PDO-forming efficiency without activating fibroblasts. Although IL-22 typically signals through the JAK–STAT pathway, it unexpectedly activates MAPK signaling in PDO cells. Interestingly, interferon-γ (IFNγ) showed only partial cytotoxic effects on CRC cells. Tumor tissues contained both IFNγ-producing ILC1 and IL-22/IL-17A–producing ILC3 populations. Co-culture with PDOs selectively induced IL-22, but not IL-17A, production in ILC3 cells. Importantly, IL-22 enhanced organoid formation only in the absence of CAFs, whereas IFNγ activity was largely unaffected by stromal context. Discussion These findings demonstrate that CAFs modulate local immunity by selectively masking ILC3-derived IL-22 signaling while preserving ILC1-mediated IFNγ responses. This study emphasizes the importance of stromal context in interpreting cytokine function in CRC and reveals a previously unrecognized mechanism of stromal–immune crosstalk within the TME.