Association of CD8+TILs co-expressing granzyme A and interferon-γ with colon cancer cells in the tumor microenvironment

Abstract CD8 + T cells secreting granzyme A (GZMA) can induce pyroptosis in tumor cells by effectively cleaving gasdermin B (GSDMB), which is stimulated by interferon-γ (IFN-γ). However, the interaction between GZMA-expressing CD8 + T cells and GSDMB-expressing tumor cells in colon cancer remains poorly understood. Our research employed multi-color immunohistochemistry (mIHC) staining and integrated clinical data to explore the spatial distribution and clinical relevance of GZMA- and IFN-γ-expressing CD8 + tumor-infiltrating lymphocytes (TILs), as well as GSDMB-expressing CK + cells, within the tumor microenvironment (TME) of human colon cancer samples. Additionally, we utilizing single-cell RNA sequencing (scRNA-seq) data to examine the functional dynamics and interactions among these cell populations. scRNA-seq analysis of colorectal cancer (CRC) tissues revealed that CD8 + TILs co-expressed GZMA and IFN-γ, but not other cell types. Our mIHC staining results indicated that a significant reduction in the infiltration of GZMA + IFN-γ + CD8 + TILs in colon cancer patients ( P < 0.01). Functional analysis results indicated that GZMA + IFN-γ + CD8 + TILs demonstrated enhanced activation and effector functions compared to other CD8 + TIL subsets. Furthermore, GSDMB-expressing CK + cells exhibited augmented immunogenicity. Correlation analysis highlighted a positive association between GSDMB + CK + cells and GZMA + IFN-γ + CD8 + TILs ( r = 0.221, P = 0.033). Analysis of cell-cell interactions further showed that these interactions were mediated by IFN-γ and transforming growth factor-β (TGF-β), the co-stimulatory molecule ICOS, and immune checkpoint molecules TIGIT and TIM-3. These findings suggested that GZMA + IFN-γ + CD8 + TILs modulating GSDMB-expressing tumor cells, significantly impacted the immune microenvironment and patients’ prognosis in colon cancer. By elucidating these mechanisms, our present study aims to provide novel insights for the advancement of immunotherapeutic strategies in colon cancer.