Decoding innate lymphoid cell heterogeneity and plasticity in colorectal cancer

BACKGROUND: In colorectal cancer (CRC), innate lymphoid cells (ILCs) play a vital role in preserving and modulating immune homeostasis within the intestinal environment. However, the origins and diverse functions of ILCs in CRC remain poorly understood, making it difficult to clarify how these cells contribute to disease progression and influence therapeutic efficacy. METHODS: Single-cell RNA sequencing (scRNA-seq) generated an atlas of ILCs from multiple tissues (bone marrow, blood, and intestine), revealing their origins, heterogeneity, and plasticity. Spatial transcriptomics (ST) and immunofluorescence (IF) defined their specific cellular neighbourhoods within the tumour microenvironment. In vitro co-culture assays were performed to validate the regulatory role of ILC2s in B cell maturation. Bulk RNA sequencing and flow cytometry were employed to assess the survival and therapeutic response potential of ILCs. RESULTS: Intestinal ILCs have two distinct origins: ILC3-CD83 cells derived from the fetal gut, which persist into adulthood; and ILC2 and ILC3-S100A4 cells that might originate from the bone marrow and migrate through the circulation to colonise intestinal tissues. The tissue-resident ILC3 subsets exhibited diverse functional roles in CRC. Specifically, trajectory analysis showed that ILC3s differentiated into either stress-responsive ILC3-HSPA1B cells or cytotoxic ILC1/NK cells in CRC. Additionally, by using spatial transcriptomics analysis combined with functional assays, we found that bone marrow-derived ILC2s preferentially localise in tertiary lymphoid structures (TLSs), where they likely support B cell maturation. Notably, higher ILC2 abundance correlated with better clinical outcomes and greater therapeutic benefit. CONCLUSIONS: This study reveals the distinct origins and functional heterogeneity of intestinal ILC subsets in CRC. The enrichment of bone marrow-derived ILC2s in TLSs, where they likely support B cell maturation, is associated with improved prognosis and favourable immunotherapy response, which may serve as biomarkers for survival and therapeutic efficacy in CRC.