Abstract B083: Mutant KRAS cooperates with the hypoxic tumor microenvironment to drive immunosuppressive macrophages via a paracrine PGE2/GM-CSF axis
Matthew T. Cribb, Ayeisha N. Colon-Ortiz, Sahar Fattani, Nefetiti Mims, Sarah Ghalayini, Sashi Kulatilaka, Morgan Green, Tara Fujimoto, Jasper R. Chen, Tianyu Wang, Kimal I. Rajapakshe, Jennifer L. Anderson, Lin Tan, Bo Wei, Iqbal Mahmud, Philip L. Lorenzi, Andrew G. Sikora, Vincent Bernard, Natividad R. Fuentes, Cullen M. Taniguchi, Albert C. Koong, Dadi Jiang, Michael T. SpiottoAbstract
INTRODUCTION:
Hypoxia is known to drive an immunosuppressive microenvironment in pancreatic ductal adenocarcinoma (PDAC). However, the underlying immunosuppressive mechanisms remain unclear. Here, we hypothesized that hypoxia-induced factors secreted from tumor cells promoted immunosuppressive macrophages in PDAC.
METHODS:
We treated macrophages with conditioned medium (CM) from the KPC pancreatic cancer cell line under normoxic or hypoxic conditions. We assessed transcriptional signatures with RNA-seq of CM-treated macrophages or from The Cancer Genome Atlas Database. T cell proliferation and activation was measured using fluorescence cytometry. We identified metabolomic and protein changes between normoxic and hypoxic KPC CM using HPLC and protein arrays, respectively. KPC cells were injected both subcutaneously and orthotopically into C57BL/6 mice and treated with combinations of celecoxib, an anti-GM-CSF neutralizing antibody, and immune checkpoint blockade with anti-CTLA-4 and anti-PD-L1. COMET multiplex immunofluorescence was used to identify changes in tumor-infiltrating T cells and macrophages following treatment.
RESULTS:
We found that hypoxia was closely correlated with immunosuppressive macrophages in human pancreatic cancers. Through metabolomic analysis, we identified prostaglandin E2 (PGE2) as a hypoxia-induced secreted factor that drove arginase (Arg1) expression in macrophages. The co-secretion of granulocyte-macrophage colony-stimulating factor (GM-CSF), which was dependent on mutant KRAS, cooperated with PGE2 to induce macrophage Arg1 expression. PGE2 and GM-CSF induced immunosuppressive transcriptional and functional changes in macrophages that phenocopied the effects of hypoxic tumor CM. Inhibition of PGE2 and GM-CSF sensitized tumors to anti-PD-L1 and anti-CTLA-4 checkpoint blockade in preclinical PDAC models and was associated with an increased M1-like macrophage phenotype and T cell infiltration. We also showed that PGE2 activated the p38 and adenylate cyclase signaling pathways in macrophages while GM-CSF activated the STAT3 pathway, together inducing the immunosuppressive transcriptional programs observed in hypoxic CM-treated macrophages.
CONCLUSION:
Our study identified that the KRAS genotype and the hypoxic tumor microenvironment cooperated to induce an immunosuppressive phenotype in macrophages via PGE2 and GM-CSF. PGE2 production was primarily dependent on hypoxia and GM-CSF secretion was dependent on mutant KRAS. Combined inhibition of PGE2 and GM-CSF sensitized PDAC tumors to checkpoint blockade, revealing a therapeutic strategy to reverse the immunosuppressive PDAC microenvironment. Overall, our work has revealed a mechanism of hypoxic crosstalk between tumor cells and macrophages which contributes to the immunologically cold PDAC microenvironment and promotes resistance to immunotherapy.
Citation Format:
Matthew T. Cribb, Ayeisha N. Colon-Ortiz, Sahar Fattani, Nefetiti Mims, Sarah Ghalayini, Sashi Kulatilaka, Morgan Green, Tara Fujimoto, Jasper R. Chen, Tianyu Wang, Kimal I. Rajapakshe, Jennifer L. Anderson, Lin Tan, Bo Wei, Iqbal Mahmud, Philip L. Lorenzi, Andrew G. Sikora, Vincent Bernard, Natividad R. Fuentes, Cullen M. Taniguchi, Albert C. Koong, Dadi Jiang, Michael T. Spiotto. Mutant KRAS cooperates with the hypoxic tumor microenvironment to drive immunosuppressive macrophages via a paracrine PGE2/GM-CSF axis [abstract]. In: Proceedings of the AACR Conference on Pancreatic Cancer: New Frontiers in Biology and Therapeutic Development; 2026 Sep 25-28; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(18_Suppl_2):Abstract nr B083.