Abstract PDAC is the most hypovascular solid tumor, imposing harsh constraints on tumor metabolism. Cancer cells cope with poor access to vascular-derived nutrients and O2 by cooperating with abundant stromal cells, receiving alanine, proline, glycosaminoglycans, branched chain ketoacids, lysophosphatidic acid, and unsaturated fatty acids from fibroblasts. However, the extent of metabolic support provided by other cell types in the PDAC TME is unexplored. We reasoned that metabolically cooperative cell types would be spatially positioned near cancer cells in these tumors, and that cooperative spatial arrangements would be enriched under conditions of relative hypovascularity. We first performed Xenium spatial transcriptomics on a cohort of 8 treatment-naive human PDAC resections and generated a spatial atlas of nearly 700, 000 cells across a range of expected cell types. Niche analysis uncovered 17 recurrent cellular neighborhoods in the data, including an endothelial-enriched niche. We find that tumor-associated neutrophils form dense aggregates within malignant glands in tumors with low endothelial content, and form in tumor regions which are most distal from vasculature. In a separate cohort of 4 additional PDAC tumors, the expression of neutrophil chemokines CXCL1/2/3/5 was highest in a subpopulation of malignant cells distant from endothelium. These data suggest metabolic stress secondary to poor vascularity drives recruitment of neutrophils via the CXCL1/2/3/5 axis. Indeed, when exposed to Tumor Interstitial Fluid Media (TIFM), cancer cells upregulated the transcription and/or secretion of these cytokines and G-CSF. Since the formation of neutrophil aggregates is driven by cancer cell nutrient derangement, we hypothesized that cancer cells could cooperate metabolically with neutrophils to alleviate nutrient stress driven by poor vascularity. Neutrophil co-culture promoted cancer cell proliferation under TIFM, but not RPMI, while under lethal combination TIFM and serum starvation, co-culture enabled enhanced survival of cancer cells. Lastly, to understand what metabolic processes could be fueled by neutrophil aggregates in pancreatic cancer, we designed a fully custom spatial transcriptomics panel targeting ∼360 metabolic enzymes and profiled 4 additional PDAC cases, performing dispersed whole-transcriptome scFFPEseq on an adjacent tissue section. We integrated data across the spatial and dispersed modalities to enable spatial projection of whole-transcriptome-derived information. Cancer cells located near neutrophil aggregates had significantly enhanced expression of vitamin B6 salvage pathway, suggesting that one critical shared metabolite between neutrophils and cancer cells is the B6 precursor pyridoxine. In total, we have deployed multiple spatial and single cell RNA profiling technologies to uncover a novel node of metabolic crosstalk in pancreatic cancer between neutrophils and cancer cells. Future work is needed to confirm transfer of specific metabolites and the downstream sequelae of their provision for tumor growth. Citation Format: Carson D. Poltorack, Connor J. Hennessey, Sydney M. Shaffer, M Celeste. Simon. Spatial profiling reveals cancer cell metabolic cooperation with aggregative neutrophils in pancreatic cancer abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research—Emerging Science Driving Transformative Solutions; Boston, MA; 2025 Sep 28-Oct 1; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2025;85 (18Suppl₃): Abstract nr B127.
Poltorack et al. (Sun,) studied this question.