Abstract 1510: Immune regulators identified in undifferentiated pleomorphic sarcoma (UPS) subpopulations

Introduction: UPS is a common subtype of soft tissue sarcoma with limited effective systemic therapies. We previously showed that a murine model of UPS has various subpopulations (SPs), characterized by distinct morphology, molecular changes, and oncogenic behaviors that likely contribute to the overall behavior of UPS. Here, we aimed to characterize interactions between UPS SPs to identify pathways important in UPS progression. Methods: We used a murine model of UPS that recapitulates the human disease. Cre recombinase drives oncogenic KRAS-G12D, P53 deletion, and expression of confetti fluorescent protein to allow for in vivo lineage tracing. Individual SPs were isolated via FACS and cultured in the presence of conditioned media (CM) from other individual SPs. Growth rate was assayed using the IncuCyte system (Essen BioScience), and cell proliferation was measured using the Click-iT EdU flow cytometry assay kit. Invasion and migration were assayed using a cell culture insert with or without Matrigel, with CM or non-CM in the lower chamber. Cells migrating/invading into the lower chamber were counted. We performed proteomics analysis on the CM from the SP that potentiated growth and invasion to define the secretome of this SP. The CCL2/CCL7/CCL8 inhibitor, bindarit, was added to CM and cell confluency was assessed as above. RNAseq data from The Cancer Genome Atlas (TCGA) were used to generate immune infiltration scores in human UPS samples. These scores were correlated with expression of lead candidates from our secretome analysis. Results: CM from one UPS SP (SP-1) increased the growth rate and invasion/migration of one other SP (SP-2). Proteomics analysis identified 233 secreted proteins in the CM from SP-1, including 37 with two-fold higher abundance compared to SP-2. Among these, several immunoregulatory proteins were identified, including serpine 1, CCL2, CCL7, CXCL1, and CX3CL1. Culture with bindarit abolished the effect of SP-1 CM on growth rate. Additionally, we found several semaphorins in the secretomes of SP-1 and SP-2, and emerging evidence suggests a key role for semaphorins in cancer progression and immune regulation. Semaphorin expression correlated with immune infiltration scores in human UPS samples. Conclusions: We identified a UPS SP that positively regulates oncogenic behaviors of other SPs through secreted factors. Additionally, several of these secreted proteins play key roles in immune regulation. Given promising activity of immunotherapy in UPS, these proteins warrant further investigation. Citation Format: Candace Lynn Haddox, Junjie Bao, Tomasa Barrientos, Benjamin A. Alman. Immune regulators identified in undifferentiated pleomorphic sarcoma (UPS) subpopulations [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1510.