Blockage of cuproplasia inhibits pancreatic tumour-associated neutrophils infiltration through TRAF6/STAT3/CCL2 pathway

Background: Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer with a poor prognosis and is easy to developing drug resistance to conventional therapies due to its distinctive tumour microenvironment. Recent advancements have brought attention to the aberrant copper metabolism in this malignancy, but the influence of intracellular cuproplasia and balance on the tumoral immune microenvironment is still uncertain. Methods: We analysed copper concentrations and CTR1 expression in PDAC tissues and cell lines. Spatial transcriptomics was employed to delineate the relationship between CTR1 overexpression and tumour-associated neutrophils (TANs) infiltration. Chemokine arrays and molecular assays were used to identify key signalling pathways involved. Functional experiments, including CTR1 knockdown and TRAF6 overexpression, were conducted to assess its impact on neutrophil infiltration and therapeutic synergy with gemcitabine. Results: We identified that CTR1 overexpression drives intracellular copper overaccumulation, activating TRAF6-dependent phosphorylation of JAK/STAT3. Phosphorylated STAT3 transcriptionally upregulates the chemokine CCL2, fostering CCR2-mediated TANs infiltration, which correlates with poor prognosis. Crucially, single-cell RNA sequencing revealed CTR1 knockdown suppresses a pro-metastatic TAN subpopulation (TAN-2) and dramatically reduces TANs recruitment in orthotopic tumour models. This copper-targeted intervention concurrently enhances cytotoxic CD8⁺ effector T cells within the TME. The translational impact is underscored in gemcitabine-resistant PDAC, where hyperactive CTR1 and intensified TANs infiltration create a therapy-refractory condition. Combining CTR1 inhibition with gemcitabine synergistically overcomes this resistance by dual remodelling of the TME. Conclusions: Our findings shed light on how intracellular copper metabolism-regulating molecules modulate the neutrophil infiltration through the TRAF6/STAT3/CCL2 pathway, and particularly, targeting the copper regulator shows potential in optimising the tumour microenvironment in the treatment of pancreatic cancers. (Figure presented.)