Purpose/Objective(s) Immunosuppressive myeloid cells in prostate cancer (PCa) are associated with disease progression and treatment resistance. Tumor-associated macrophages (TAMs) contribute to progression through suppression of anti-tumor immunity, remodeling of the tumor microenvironment (TME) and promoting tumor cell invasion and metastasis. SBRT is increasingly used for treatment of localized and metastatic PCa and promotes inflammatory remodeling of the TME. We examined irradiated human PCa in a cohort of patients that underwent pre-operative SBRT followed by prostatectomy (NCT03663218) to identify radiation-induced targets and validated findings in both in vitro and in vivo models. Materials/Methods We performed 10X single-cell RNA sequencing (scRNA-seq) on prostatectomy specimens from men with high-risk localized PCa; 12,988 cells from 4 patients that underwent pre-operative SBRT and 47,314 cells from 6 patients that underwent prostatectomy alone. THP-1 monocytes polarized into macrophage states (M0, M1, M2, TAM) were used to characterize TREM2 expression and immune responses to radiation. In vivo studies with C57BL/6 mice inoculated with B6CaP flank tumors or PTEN-/-SPOPmutCHD1del organoid cell line were used to analyze intratumoral myeloid populations in response to SBRT (37.5Gy in 5 fractions). Results scRNA-seq analysis (Cell Ranger 7.1.0; Seurat v5 R package) revealed enrichment of a cluster of myeloid cells defined by a damage-associated Mɸ gene signature (TREM2, APOE, SPP1, LGALS3, CD9) in response to SBRT compared with non-irradiated specimens (35.6% vs 5.7%). Irradiation (20Gy x1) of THP-1 cells significantly increased TREM2 expression in M0 (p<0.01), M1 (p<0.001), and TAM (p<0.01) states. Conditioned media from irradiated THP-1 TAMs increased proliferation of PC3 tumor cells (p<0.0001) whereas CM (p<0.05) and protein lysate (p<0.001) derived from irradiated PC3 cells significantly increased TREM2 expression on M0-polarized THP-1 cells. In vivo studies with prostate SBRT in the organoid and B6CaP model both demonstrated significant irradiated tumor control (p<0.0001). TREM2 expression and sTREM2 levels were significantly increased in the irradiated TME (p<0.05) in both models as well as SPP1 expression in the B6CaP model. Immune profiling demonstrated robust myeloid infiltration in response to SBRT dominated by CD11b+F4/80+ TAMs in irradiated tumors. Furthermore, among irradiated tumors, TREM2 expression was markedly elevated in TAMs (organoid, p<0.0001) and TREM2 mean fluorescence intensity was significantly enriched among CD206+ TAMs in organoid (p< 0.001) and B6CaP (p<0.01) models. Conclusion Preliminary findings in irradiated human PCa and subsequent validation studies in preclinical models suggest TREM2 defines a subset of radiation-induced TAMs. Ongoing work aims to characterize functional implications of radiation-induced TREM2 expression and whether TREM2 can be targeted to modulate radiation-induced immune responses.

Radiation-Induced TREM2-Expression on Tumor-Associated Macrophages is a Target for Modulation of Immune Responses in Prostate Cancer