Abstract 3979: Genomic determinants of canine tumor-associated macrophage polarization

Abstract There is poorly understood variability both between and within cancer types in their ability to maintain an immunosuppressive microenvironment and polarize tumor-associated macrophages (TAMs). Identifying genetic biomarkers of cancers that strongly polarize TAMs would enable the discovery of novel therapeutic targets and support the stratification of patients for appropriate immunotherapy. Our aim, therefore, was to correlate gene expression data from cancer cell lines with phenotypic changes in canine macrophages. We harvested standardized tumor-conditioned medium (TCM) from twenty-five canine cancer cell lines with bulk RNA-sequencing data available. Primary monocyte-derived macrophages were obtained from three canine donors and polarized with the twenty-five different TCMs. After a washout period, supernatants were collected, and polarization changes of the macrophages were assessed via secretory products using ELISA and a multiplex cytokine/chemokine assay. For each analyte measured, a modified z-score was used to rank each cell line’s ability to stimulate the secretion of that analyte from macrophages. Multiple regression analysis and pairwise comparison analysis (composed of “strong” and “weak” stimulators) were performed against the full transcriptome. Many significantly differentially expressed genes were identified in the cancer cell lines with the strongest ability to polarize macrophages. For example, cell lines with MVB12A upregulated, a gene involved in selecting cargo for exosomes, had the highest ability to stimulate VEGF production from macrophages. This was subsequently confirmed by depleting the TCM of exosomes which resulted in a significant decrease in VEGF production. Multiple other genes involved in macrophage immunosuppression were upregulated, while those involved in cell adhesion and metastasis prevention were downregulated. In conclusion, canine cancer cell lines have varying abilities to polarize primary macrophages, irrespective of histologic type, as measured by secretory products. The cell lines capable of potent polarization had relevant differentially expressed genes and pathways. These are currently being explored to identify novel biomarkers and therapeutic targets. Citation Format: Rachel V. Brady, Kristen B. Farrell, Eric P. Palmer, Sunetra Das, Dawn L. Duval, Douglas H. Thamm. Genomic determinants of canine tumor-associated macrophage polarization [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3979.