Disruption of ACTG1 potentiates anti-TIM-3 immunotherapy by reprogramming the immunosuppressive microenvironment via the CSF3/CSF3R axis in pancreatic cancer

Abstract

Background The limited immune infiltration observed in pancreatic ductal adenocarcinoma (PDAC) is a major factor contributing to immunotherapy resistance and the consistently poor prognosis. Hence, identifying novel therapeutic targets and developing effective combination strategies to overcome immunotherapy resistance represent urgent and unmet clinical needs. Methods The gene expression datasets and corresponding clinical information were collected from TCGA, GEO and ICGC. The immune cell infiltration (ICI) contents were evaluated by “CIBERSORT”. PDAC subtypes based on ICI pattern were obtained from consensus clustering. Their prognostic value was validated by the Kaplan-Meier survival curves. Then, we employed an integrated multi-dataset approach to systematically identify pivotal regulators of PDAC progression. The functional role of ACTG1 was specifically validated through in vivo and in vitro models. The tumor immune microenvironment was characterized by flow cytometry and immunohistochemistry (IHC). Potential molecular mechanisms were further investigated using RNA sequencing and tumor cell-macrophage co-culture experiments. Results Three distinct immune infiltration subtypes with different prognoses and immunological features were identified in the 608-sample GEO/ICGC training cohort and the 177-sample TCGA-PAAD validation cohort. The ICI pattern with a better prognosis was characterized by high level of plasma cells, activated NK cells, activated DC cells, M1 macrophages and CD8+ T cells, low level of M0 macrophages, M2 macrophages. ACTG1 drives immunosuppression and predicts poor prognosis in PDAC. Mechanistically, ACTG1 promotes tumor cell-derived CSF3 secretion, which upregulates TIM-3 expression on tumor-associated macrophages (TAMs) via the CSF3/CSF3R signaling pathway. This cascade reinforces M2-like TAMs polarization and induces CD8+ T cell exhaustion, thereby fostering an immunosuppressive niche. Strikingly, ACTG1 knockdown combined with TIM-3 blockade exerts an enhanced effect in reversing immunosuppression, restoring anti-tumor immunity, and markedly prolonging survival. Conclusions ACTG1 promotes PDAC progression by orchestrating an immunosuppressive tumor microenvironment through the CSF3/CSF3R/TIM-3 axis. Thus, combining genetic suppression of ACTG1 with TIM-3 blockade represents a potent therapeutic approach to overcome immune evasion and curb tumor progression.