Tumor microenvironment is characterized by chronic inflammation represented by infiltrating leukocytes and soluble mediators, which lead to a local and systemic immunosuppression associated with cancer progression. Here, we used the ret transgenic spontaneous murine melanoma model that mimics human melanoma. Skin tumors and metastatic lymph nodes showed increased levels of inflammatory factors such as IL-1β, GM-CSF, and IFN-γ, which correlated with tumor progression. Moreover, Gr1 + CD11b + myeloidderived suppressor cells (MDSCs), known to inhibit tumor reactive T cells, were enriched in melanoma lesions and lymphatic organs during tumor progression. MDSC infiltration was associated with a strong TCR ζ-chain down-regulation in all T cells. Coculturing normal splenocytes with tumor-derived MDSC induced a decreased T-cell proliferation and ζ-chain expression, verifying the MDSC immunosuppressive function and suggesting that the tumor inflammatory microenvironment supports MDSC recruitment and immunosuppressive activity. Indeed, upon manipulation of the melanoma microenvironment with the phosphodiesterase-5 inhibitor sildenafil, we observed reduced levels of numerous inflammatory mediators (e.g., IL-1β, IL-6, VEGF, S100A9) in association with decreased MDSC amounts and immunosuppressive function, indicating an antiinflammatory effect of sildenafil. This led to a partial restoration of ζ-chain expression in T cells and to a significantly increased survival of tumor-bearing mice. CD8 T-cell depletion resulted in an abrogation of sildenafil beneficial outcome, suggesting the involvement of MDSC and CD8 T cells in the observed therapeutic effects. Our data imply that inhibition of chronic inflammation in the tumor microenvironment should be applied in conjunction with melanoma immunotherapies to increase their efficacy.therapy | cytokines

(2015) Extracellular vesicle-mediated transfer of functional RNA in the tumor microenvironment, OncoImmunology, 4:6, e1008371,

Purpose: Immunotherapy with ipilimumab improves the survival of patients with metastatic melanoma. Because only around 20% of patients experience long-term benefit, reliable markers are needed to predict a clinical response. Therefore, we sought to determine if some myeloid cells and related inflammatory mediators could serve as predictive factors for the patients' response to ipilimumab.Experimental Design: We performed an analysis of myeloid cells in the peripheral blood of 59 stage IV melanoma patients before the treatment and at different time points upon the therapy using a clinical laboratory analysis and multicolor flow cytometry. In addition, the production of related inflammatory factors was evaluated by ELISA or Bio-Plex assays.Results: An early increase in eosinophil count during the treatment with ipilimumab was associated with an improved clinical response. In contrast, elevated amounts of monocytic myeloid-derived suppressor cells (moMDSC), neutrophils, and monocytes were found in nonresponders (n ¼ 36) as compared with basal levels and with responding patients (n ¼ 23). Moreover, in nonresponders, moMDSCs produced significantly more nitric oxide, and granulocytic MDSCs expressed higher levels of PD-L1 than these parameters at baseline and in responders, suggesting their enhanced immunosuppressive capacity. Upon the first ipilimumab infusion, nonresponders displayed elevated serum concentrations of S100A8/A9 and HMGB1 that attract and activate MDSCs.Conclusions: These findings highlight additional mechanisms of ipilimumab effects and suggest levels of eosinophils, MDSCs, as well as related inflammatory factors S100A8/A9 and HMGB1 as novel complex predictive markers for patients who may benefit from the ipilimumab therapy. Clin Cancer Res; 21(24); 5453-9. Ó2015 AACR.

SummaryTumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is known for specifically killing cancer cells, whereas in resistant cancers, TRAIL/TRAIL-R can promote metastasis via Rac1 and PI3K. It remains unknown, however, whether and to what extent TRAIL/TRAIL-R signaling in cancer cells can affect the immune microenvironment. Here we show that TRAIL-triggered cytokine secretion from TRAIL-resistant cancer cells is FADD dependent and identify the TRAIL-induced secretome to drive monocyte polarization to myeloid-derived suppressor cells (MDSCs) and M2-like macrophages. TRAIL-R suppression in tumor cells impaired CCL2 production and diminished both lung MDSC presence and tumor growth. In accordance, the receptor of CCL2, CCR2, is required to facilitate increased MDSC presence and tumor growth. Finally, TRAIL and CCL2 are co-regulated with MDSC/M2 markers in lung adenocarcinoma patients. Collectively, endogenous TRAIL/TRAIL-R-mediated CCL2 secretion promotes accumulation of tumor-supportive immune cells in the cancer microenvironment, thereby revealing a tumor-supportive immune-modulatory role of the TRAIL/TRAIL-R system in cancer biology.

The anti-tumor effects of paclitaxel are generally attributed to the suppression of microtubule dynamics resulting in defects in cell division. New data demonstrated that in ultra-low non-cytotoxic concentrations, paclitaxel modulated in immune cells in vitro the activity of small Rho GTPases, the key regulators of intracellular actin dynamics. However, the immunomodulatory properties of paclitaxel in vivo have not been evaluated. Using here the ret transgenic murine melanoma model, which mimics human cutaneous melanoma, we tested effects of ultra-low non-cytotoxic dose paclitaxel on functions of myeloid-derived suppressor cells (MDSCs), chronic inflammatory mediators, and T cell activities in the tumor microenvironment in vivo. Administration of paclitaxel significantly decreased accumulation and immunosuppressive activities of tumor-infiltrating MDSCs without alterations of the bone marrow hematopoiesis. This was associated with the inhibition of p38 MAPK activity, TNF-α and production and S100A9 expression in MDSCs. The production of mediators of chronic inflammation in the tumor milieu was also diminished. Importantly, reduced tumor burden and increased animal survival upon paclitaxel application was mediated by the restoration of CD8 T cell effector functions. We suggest that the ability of paclitaxel in non-cytotoxic dose to block the immunosuppressive potential of MDSCs in vivo represents a new therapeutic strategy to down-regulate immunosuppression and chronic inflammation in the tumor microenvironment for enhancing the efficacy of concomitant anti-cancer therapies.

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Myeloid cells play a key role in the outcome of anti-tumor immunity and response to anti-cancer therapy, since in the tumor microenvironment they may exert both stimulatory and inhibitory pressures on the proliferative, angiogenic, metastatic, and immunomodulating potential of tumor cells. Therefore, understanding the mechanisms of myeloid regulatory cell differentiation is critical for developing strategies for the therapeutic reversal of myeloid derived suppressor cell (MDSC) accumulation in the tumor-bearing hosts. Here, using an in vitro model system, several potential mechanisms of the direct effect of paclitaxel on MDSC were tested, which might be responsible for the anti-tumor potential of low-dose paclitaxel therapy in mice. It was hypothesized that a decreased level of MDSC in vivo after paclitaxel administration might be due to (i) the blockage of MDSC generation, (ii) an induction of MDSC apoptosis, or (iii) the stimulation of MDSC differentiation. The results revealed that paclitaxel in ultra-low concentrations neither increased MDSC apoptosis nor blocked MDSC generation, but stimulated MDSC differentiation towards dendritic cells. This effect of paclitaxel was TLR4-independent since it was not diminished in cell cultures originated from TLR4−/− mice. These results support a new concept that certain chemotherapeutic agents in ultra-low non-cytotoxic doses may suppress tumor progression by targeting several cell populations in the tumor microenvironment, including MDSC.

Tumor progression is often associated with chronic inflammation in the tumor microenvironment, which is mediated by numerous cytokines, chemokines and growth factors produced by cancer and stroma cells. All these mediators support tumor development and immunosuppression in autocrine and/or paracrine ways. Neutralization of chronic inflammatory conditions can lead to the restoration of anti-tumor immune responses. Among stroma cells infiltrating tumors, myeloid-derived suppressor cells (MDSCs) represent one of the most important players mediating immunosuppression. These cells may not only inhibit an anti-tumor immunity but also directly stimulate tumorigenesis as well as tumor growth and expansion. Therefore, understanding the mechanisms of generation, migration to the tumor site and activation of MDSC is necessary for the development of new strategies of tumor immunotherapy.