NSUN6 deficiency drives immune suppression in pancreatic cancer via the KDM5A-CCL2-macrophage axis

Background RNA 5-methylcytosine (m 5 C) has emerged as a critical epigenetic regulator in cancer biology, yet its role in the tumour immune microenvironment (TME) remains incompletely understood. Objective We aimed to elucidate the functional role and underlying mechanism of NOP2/Sun RNA methyltransferase 6 (NSUN6), an m 5 C methyltransferase, in shaping the TME of pancreatic ductal adenocarcinoma (PDAC). Design The clinical significance of NSUN6 was assessed in human PDAC cohorts. The impact of NSUN6 on antitumour immunity was evaluated using murine PDAC models. Single-cell RNA sequencing was employed to characterise the TME landscape of PDAC. RNA bisulfite sequencing and RNA sequencing were used to identify NSUN6 targets. The synergistic effects of C-C motif chemokine ligand 2 (CCL2) blockade combined with immune checkpoint blockade (ICB) therapy were investigated. Results NSUN6 deficiency significantly enhanced macrophage accumulation and polarisation toward immunosuppressive phenotypes, thereby impairing CD8 + T cell-mediated antitumour immunity in PDAC. Mechanistically, NSUN6 deficiency downregulated KDM5A expression in an m 5 C-dependent manner, resulting in transcriptional activation of CCL2. Elevated CCL2 secretion promoted the accumulation and polarisation of protumorous macrophages, fostering an immunosuppressive TME and inducing resistance to ICB. Notably, CCL2 blockade reversed protumorous macrophage infiltration and restored ICB sensitivity in Nsun6-deficient murine PDAC models. Clinical analysis further revealed a positive correlation between NSUN6 expression and favourable immune responses in ICB-treated cohorts. Conclusion NSUN6 deficiency drives immune suppression through the m 5 C-KDM5A-CCL2 axis in PDAC. Targeting the NSUN6-CCL2 axis represents a promising strategy to sensitise PDAC to ICB therapy.