IL-27 shapes NK cell heterogeneity and function in colorectal cancer

Colorectal cancer (CRC) is a leading cause of cancer-related mortality worldwide and is characterized by an immunosuppressive tumor microenvironment (TME). While adaptive immunity contributes to tumor control, growing evidence underscores the role of innate lymphocytes, particularly natural killer (NK) cells, in early antitumor surveillance. However, tumor-infiltrating NK cells often exhibit defective maturation and impaired effector functions, whereas the signals regulating NK cell differentiation and activity in CRC remain poorly defined. Interleukin-27 (IL-27) has emerged as a regulator of antitumor immunity, yet its role in modulating NK cell responses in intestinal tumors is largely unexplored.

We employed an orthotopic transplantation model of genetically engineered colorectal tumor organoids Apc^-/-Kras^G12D/+Trp53^R172H/-Smad4^-/- (AKPS) to investigate NK cell heterogeneity, maturation, and function during CRC progression. Tumor-infiltrating innate lymphocytes were analyzed using single-cell RNA sequencing, flow cytometry, immunofluorescence, and functional assays. In vitro co-culture systems and in vivo IL-27 blockade were used to assess the impact of tumor-derived IL-27 on NK cell transcriptional programs and effector activity.

Single-cell transcriptomic profiling revealed marked heterogeneity among tumor-infiltrating NK cells, identifying subsets with different maturation states and functional capacities. Among innate lymphocytes, AKPS tumors were dominated by NK cells displaying reduced activating receptors and diminished cytotoxic and cytokine-producing potential. Phenotypic and adoptive transfer analyses demonstrated that the TME favors persistence of immature CD27⁺ NK cell subsets while limiting differentiated NK cells. Mechanistically, tumor-derived IL-27 emerged as a critical regulator sustaining NK cell infiltration and activation; IL-27 blockade reduced NK cell accumulation, interferon-γ production, and immature subset frequency. The clinical relevance of these findings was supported by analysis of human CRC single-cell datasets, which revealed elevated IL-27 signaling in intratumoral NK cells, as well as by the ability of patient-derived organoids to enhance NK-cell cytotoxicity, which was reduced by IL-27 blockade.

Our study identifies IL-27 as a critical modulator of NK cell differentiation and function in CRC, highlighting its role in sustaining NK cell-mediated immune surveillance within the tumor microenvironment. These findings provide mechanistic insight into NK cell dysfunction in CRC and suggest IL-27 signaling as a promising therapeutic target to restore innate antitumor immunity.