Pancreatic ductal adenocarcinoma (PDAC) is resistant to most therapies including single-agent immunotherapy and has a dense desmoplastic stroma, and most patients present with advanced metastatic disease. We reveal that macrophages are the dominant leukocyte population both in human PDAC stroma and autochthonous models, with an important functional contribution to the squamous subtype of human PDAC. We targeted macrophages in a genetic PDAC model using AZD7507, a potent selective inhibitor of CSF1R. AZD7507 caused shrinkage of established tumors and increased mouse survival in this difficult-to-treat model. Malignant cell proliferation diminished, with increased cell death and an enhanced T cell immune response. Loss of macrophages rewired other features of the TME, with global changes in gene expression akin to switching PDAC subtypes. These changes were markedly different to those elicited when neutrophils were targeted via CXCR2. These results suggest targeting the myeloid cell axis may be particularly efficacious in PDAC, especially with CSF1R inhibitors. Display omitted •Macrophages functionally contribute to the squamous subtype of human PDAC•Inhibition of CSF1R alters the TME and results in an enhanced T cell immune response•Loss of macrophages rewires PDAC gene expression and switches subtype•Macrophage inhibition induces changes markedly different to neutrophil targeting Candido et al. find that CSF-1R+ macrophages play a vital role in pancreatic tumor maintenance, suppression of T cells, and tumor gene networks based on transcriptome analysis. In an autochthonous model, CSF-1R inhibition alters the gene expression programs that influence subtype specification of PDAC and extends survival.