Titolo della tesi: Modeling colorectal liver metastasis to leverage type I interferon-induced innate immune cell anti-tumor response
Introduction
Colorectal cancer liver metastasis (CRLM) is the leading cause of death in patients with colorectal cancer, and its immune landscape is profoundly shaped by tumor-intrinsic pathways. Among the key regulators of liver immunity, type 1 innate lymphoid cells (ILC1), natural killer (NK) cells, and metastasis-associated macrophages (MAMs) orchestrate tissue residency, effector activation, and immunoregulation. However, the features of the tumor microenvironment, particularly those associated with mismatch-repair-deficient (dMMR) and proficient (pMMR) tumors, shape the recruitment and functional polarization of these populations remain poorly understood.
Aim
This study investigates how tumors with distinct genetic features and type I interferon programs regulate the immune architecture of CRLM. By comparing two orthotopic metastasis models, MC38 (dMMR, immunogenic) and SL4 (pMMR, non-immunogenic), we dissect mechanisms controlling NK/ILC1 recruitment, differentiation, and interaction with MAMs, and assess whether STING pathway activation can convert poorly immunogenic lesions into immune-responsive metastases.
Results and Methods
Using flow cytometry, transcriptional analysis, conditional CXCR3 deletion in MC38-derived NKp46+ cells, macrophage and lymphocyte co-cultures, and in vivo treatments, we uncovered pronounced differences between the two CRC models.
MC38 metastases displayed abundant, functionally competent type 1 ILC/NK populations, including strongly infiltrating CD49a⁺ tissue-resident NK cells with mature, cytotoxic profiles (CD69, CD11b, high effector activity). SL4 lesions instead showed limited NK recruitment, reduced maturation, and weaker effector responses.
Loss of CXCR3 impaired MC38-derived NK infiltration, reduced acquisition of residency features, and increased metastatic burden, confirming CXCR3 as a central determinant of MC38-derived NK access to the metastatic niche.
Myeloid profiling revealed that MC38 metastases were enriched in mature, CXCL9-producing MAMs with a type I IFN-driven program, whereas SL4 MAMs expressed ECM-remodeling and IFN--associated signatures. Differences of macrophage polarization resulted in a different capacity to shape NK cell function in co-culture experiments: MC38-derived MAMs promoted stronger NK activation and IFN- production compared to SL4 MAMs.
Both MC38 tumor cells and MAMs exhibited a more active baseline I IFN pathway.
In vivo, diABZI treatment activated type I IFN programs in both models, promoting lymphocyte activation, phenotypic remodeling of lymphoid and myeloid cells, and a marked increase in PD-L1 expression, with especially strong induction on CD45- tumor cells in SL4. This observation prompted testing of whether STING activation could sensitize poorly immunogenic SL4 metastases to immune checkpoint blockade.
Whereas anti-PD-L1 alone was effective only in MC38, combining diABZI with anti-PD-L1 in SL4 profoundly reduced metastatic burden. This response was accompanied by expansion of proliferative, tissue-resident CD8⁺ T cells, activation of NK/ILC1 subsets, and a shift toward less immunosuppressive macrophage states.
Conclusions
Tumor-intrinsic type I IFN programs strongly associate with the immune organization of colorectal liver metastases by regulating NK recruitment, differentiation, and function through CXCR3-CXCL9 interactions and macrophage-derived cues. Enhancing STING signaling can convert immunologically cold, pMMR-like metastases into lesions responsive to PD-L1 blockade. These findings support STING-based combination strategies as a promising therapeutic avenue for poorly immunogenic CRLM.