Tumor-associated stromal myofibroblasts are essential for the progression and metastatic spread of solid tumors. Corresponding myeloid cell infiltration into primary tumors is a negative prognostic factor in some malignancies. The aim of this study was to define the exact role of stromal myofibroblasts and stromal factors in early prostate carcinoma (PCa) regulating monocyte infiltration and differentiation into dendritic cells (DCs). Epithelial and stromal primary cultures were generated from PCa biopsies and their purity confirmed. Stromal cells produced significantly more of the (C-C) motif chemokine ligand 2 (CCL2), interleukin 6 (IL-6) and transforming growth factor β (TGFβ) than epithelial cells. Monocyte chemoattraction was predominantly due to stromal-derived factors, mainly CCL2. DCs generated in the presence of stromal (but not epithelial) factors upregulated CD209, but failed to downregulate the monocyte marker CD14 in a signal transducer and activator of transcription 3 (STAT3)-dependent manner. Monocytes exposed to stromal factors did not produce detectable amounts of IL-10, however, upon lipopolysaccharide stimulation, stromal factor generated dendritic cells (sDC) produced significantly more IL-10 and less IL-12 than their conventional DC counterparts. sDC failed to cross-present tumor-antigen to CD8+ T cells and suppressed T-cell proliferation. Most importantly, sDC expressed significantly elevated levels of programmed cell death ligand-1 (PD-L1) in a primarily STAT3 and IL-6-dependent manner. In parallel with our findings in vitro, tumor-infiltrating CD14+ cells in situ were found to express both PD-L1 and CD209, and a higher percentage of tumor-associated CD3+ T cells expressed programmed cell death-1 (PD-1) molecules compared to T cells in blood. These results demonstrate a hitherto undescribed, fundamental contribution of tumor-associated stromal myofibroblasts to the development of an immunosuppressive microenvironment in early PCa.
Keywords: CCL2; CCL2, (C–C) motif chemokine ligand-2; CFSE, carboxyfluorescein succinimidyl ester; CK, cytokeratin; CM, conditioned media; CXCL, chemokine (C–X–C) motif; DC, dendritic cell; ELISA, enzyme-linked immunosorbent assay; GM-CSF, granulocyte macrophage colony-stimulating factor; HFF, human foreskin fibroblast; HGF, hepatocyte growth factor; I-TAC, interferon-inducible T cell α chemoattractant; IFN, interferon; IL, interleukin; IL-6; IP-10, interferon-γ induced protein 10; LPS, lipopolysaccharide; MIF, macrophage inhibitory factor; PBMC, peripheral blood mononuclear cells; PCaEp, prostate cancer epithelia; PCaSt, prostate cancer stroma; PD-1, programmed cell death-1; PD-L1; PD-L1, programmed cell death ligand-1; RANTES/CCL5, regulated on activation, normal T cell expressed and secreted; SCBM, stromal cell basal media; SDF-1, stromal-derived factor-1; STAT3; STAT3, signal transducer and activator of transcription 3; TGFβ, transforming growth factor β; TIL, tumor infiltrating leukocytes; VEGF, vascular endothelial growth factor; antigen cross-presentation; dendritic cells; immunosuppression; prostate cancer; sDC, DC generated in the presence of 50% PCaSt-CM; tumor microenvironment; tumor stroma; α-SMA, α-smooth muscle actin.