Luigi Dolcetti

Tumors induce a subset of inflammatory monocytes with immunosuppressive activity on CD8+ T cells

Active suppression of tumor-specific T lymphocytes can limit the efficacy of immune surveillance and immunotherapy. While tumor-recruited CD11b+ myeloid cells are known mediators of tumor-associated immune dysfunction, the true nature of these suppressive cells and the fine biochemical pathways governing their immunosuppressive activity remain elusive. Here we describe a population of circulating CD11b+IL-4 receptor alpha+ (CD11b+IL-4Ralpha+), inflammatory-type monocytes that is elicited by growing tumors and activated by IFN-gamma released from T lymphocytes. CD11b+IL-4Ralpha+ cells produced IL-13 and IFN-gamma and integrated the downstream signals of these cytokines to trigger the molecular pathways suppressing antigen-activated CD8+ T lymphocytes. Analogous immunosuppressive circuits were active in CD11b+ cells present within the tumor microenvironment. These suppressor cells challenge the current idea that tumor-conditioned immunosuppressive monocytes/macrophages are alternatively activated. Moreover, our data show how the inflammatory response elicited by tumors had detrimental effects on the adaptive immune system and suggest novel approaches for the treatment of tumor-induced immune dysfunctions.

Phosphodiesterase-5 inhibition augments endogenous antitumor immunity by reducing myeloid-derived suppressor cell function

Phosphodiesterase-5 (PDE5) inhibitors (sildenafil, tadalafil, and vardenafil) are agents currently in clinical use for nonmalignant conditions. We report the use of PDE5 inhibitors as modulators of the antitumor immune response. In several mouse tumor models, PDE5 inhibition reverses tumor-induced immunosuppressive mechanisms and enables a measurable antitumor immune response to be generated that substantially delays tumor progression. In particular, sildenafil, down-regulates arginase 1 and nitric oxide synthase–2 expression, thereby reducing the suppressive machinery of CD11b+/Gr-1+ myeloid-derived suppressor cells (MDSCs) recruited by growing tumors. By removing these tumor escape mechanisms, sildenafil enhances intratumoral T cell infiltration and activation, reduces tumor outgrowth, and improves the antitumor efficacy of adoptive T cell therapy. Sildenafil also restores in vitro T cell proliferation of peripheral blood mononuclear cells from multiple myeloma and head and neck cancer patients. In light of the recent data that enzymes mediating MDSC-dependent immunosuppression in mice are active also in humans, these findings demonstrate a potentially novel use of PDE5 inhibitors as adjuncts to tumor-specific immune therapy.

Tumor-Induced Tolerance and Immune Suppression Depend on the C/EBPβ Transcription Factor

Tumor growth is associated with a profound alteration in myelopoiesis, leading to recruitment of immunosuppressive cells known as myeloid-derived suppressor cells (MDSCs). We showed that among factors produced by various experimental tumors, the cytokines GM-CSF, G-CSF, and IL-6 allowed a rapid generation of MDSCs from precursors present in mouse and human bone marrow (BM). BM-MDSCs induced by GM-CSF+IL-6 possessed the highest tolerogenic activity, as revealed by the ability to impair the priming of CD8(+) T cells and allow long term acceptance of pancreatic islet allografts. Cytokines inducing MDSCs acted on a common molecular pathway and the immunoregulatory activity of both tumor-induced and BM-derived MDSCs was entirely dependent on the C/EBPbeta transcription factor. Adoptive transfer of tumor antigen-specific CD8(+) T lymphocytes resulted in therapy of established tumors only in mice lacking C/EBPbeta in the myeloid compartment, suggesting that C/EBPbeta is a critical regulator of the immunosuppressive environment created by growing cancers.

Hierarchy of immunosuppressive strength among myeloid-derived suppressor cell subsets is determined by GM-CSF

CD11b+/Gr‐1+ myeloid‐derived suppressor cells (MDSC) contribute to tumor immune evasion by restraining the activity of CD8+ T‐cells. Two major MDSC subsets were recently shown to play an equal role in MDSC‐induced immune dysfunctions: monocytic‐ and granulocytic‐like. We isolated three fractions of MDSC, i.e. CD11b+/Gr‐1high, CD11b+/Gr‐1int, and CD11b+/Gr‐1low populations that were characterized morphologically, phenotypically and functionally in different tumor models. In vitro assays showed that CD11b+/Gr‐1int cell subset, mainly comprising monocytes and myeloid precursors, was always capable to suppress CD8+ T‐cell activation, while CD11b+/Gr‐1high cells, mostly granulocytes, exerted appreciable suppression only in some tumor models and when present in high numbers. The CD11b+/Gr‐1int but not CD11b+/Gr‐1high cells were also immunosuppressive in vivo following adoptive transfer. CD11b+/Gr‐1low cells retained the immunosuppressive potential in most tumor models. Gene silencing experiments indicated that GM‐CSF was necessary to induce preferential expansion of both CD11b+/Gr‐1int and CD11b+/Gr‐1low subsets in the spleen of tumor‐bearing mice and mediate tumor‐induced tolerance whereas G‐CSF, which preferentially expanded CD11b+/Gr‐1high cells, did not create such immunosuppressive environment. GM‐CSF also acted on granulocyte–macrophage progenitors in the bone marrow inducing local expansion of CD11b+/Gr‐1low cells. These data unveil a hierarchy of immunoregulatory activity among MDSC subsets that is controlled by tumor‐released GM‐CSF.

Derangement of immune responses by myeloid suppressor cells

In tumor-bearing mice and cancer patients, tumor progression is often associated with altered hematopoiesis leading to the accumulation of myeloid cells. Extensive studies in preclinical models indicate that these cells share the CD11b and the Gr-1 markers, possess a mixed mature-immature myeloid phenotype, and are responsible for the induction of T-cell dysfunctions, both tumor-specific and nonspecific. Moreover, CD11b+Gr-1+ myeloid cells are described under different unrelated situations associated with temporary impairment of the T-lymphocyte reactivity. This review examines recent findings on the nature, properties, and mechanisms of action of these myeloid suppressor cells (MSCs).

Myeloid-derived suppressor cell role in tumor-related inflammation

Chronic inflammatory state can create a proper environment for neoplastic onset and sustain cancer growth. The inflammatory state that arises at the tumor edge could contribute to immune escape phenomena in many ways. Myeloid-derived suppressor cells (MDSCs), a cell population that contributes to tumor escape, immune tolerance, and suppression, respond to a variety of pro-inflammatory and anti-inflammatory stimuli, which drive their recruitment and activation. Understanding how the inflammatory milieu favours tumor escape through the accumulation of MDSCs could be very useful to improve the efficacy of cancer immunotherapy.

Susceptibility to genetic damage and cell types in Mediterranean mussels

Micronucleus (MN) frequency is generally accepted as a marker of chromosomal damage and has been studied in a variety of cells and species. In previous work, we detected significant dose-related MN increases in the epithelial-like gill cells and agranular haemocytes of Mytilus galloprovincialis treated with benzo[a]pyrene, a well-known mutagenic pollutant. In addition, we have studied micronuclei and other nuclear abnormalities in mussels collected from the Venice lagoon (Italy). Frequency changes, possibly related to genotoxic/toxic stress, in both granular and micronucleated cells from gills and haemolymph, were detected. Environmental data suggest the effect of genotoxic pollutants and the importance of cell replication in the interpretation of micronucleus frequencies.

Inhibition of Tumor-Induced Myeloid-Derived Suppressor Cell Function by a Nanoparticulated Adjuvant

The interaction between cancer vaccine adjuvants and myeloid-derived suppressor cells (MDSCs) is currently poorly understood. Very small size proteoliposomes (VSSP) are a nanoparticulated adjuvant under investigation in clinical trials in patients with renal carcinoma, breast cancer, prostate cancer, and cervical intraepithelial neoplasia grade III. We found that VSSP adjuvant induced a significant splenomegaly due to accumulation of CD11b+Gr-1+ cells. However, VSSP-derived MDSCs showed a reduced capacity to suppress both allogeneic and Ag-specific CTL response compared with that of tumor-induced MDSCs. Moreover, splenic MDSCs isolated from tumor-bearing mice treated with VSSP were phenotypically more similar to those isolated from VSSP-treated tumor-free mice and much less suppressive than tumor-induced MDSCs, both in vitro and in vivo. Furthermore, different from dendritic cell vaccination, inoculation of VSSP-based vaccine in EG.7-OVA tumor-bearing mice was sufficient to avoid tumor-induced tolerance and stimulate an immune response against OVA Ag, similar to that observed in tumor-free mice. This effect correlated with an accelerated differentiation of MDSCs into mature APCs that was promoted by VSSP. VSSP used as a cancer vaccine adjuvant might thus improve antitumor efficacy not only by stimulating a potent immune response against tumor Ags but also by reducing tumor-induced immunosuppression.

Leukocyte infiltration in cancer creates an unfavorable environment for antitumor immune responses: a novel target for therapeutic intervention.

The interaction between tumor cells and the nearby environment is being actively investigated to explore how this interplay affects the initiation and progression of cancer. Host-tumor relationship results in the production of pro-inflammatory cytokines and chemokines that promote the recruitment of leukocytes within and around developing neoplasms. Cancer cells, together with newly recruited tumor-infiltrating cells, can also activate fibroblast and vascular responses, thus resulting in a chronic microenvironment perturbation. In this complex scenario, interactions between innate and adaptive immune cells can be disturbed, leading to a failure of immune-mediated tumor recognition and destruction. On the basis of the recent awareness about tumor promotion and immune deregulation by immune/inflammatory cells, novel anti-cancer strategies can be exploited.

Apoptosis in mesenchymal stromal cells induces in vivo recipient-mediated immunomodulation

Mesenchymal stromal cell apoptosis, induced in vivo by recipient cytotoxic cells, is required for immunosuppression and clinical responses. MSC sacrifice for immunosuppression Transfer of mesenchymal stromal cells (MSCs) induces immunosuppression, although the cells are undetectable shortly after transfer. The immunosuppressive mechanism of MSCs has been somewhat of a mystery, but Galleu and colleagues now suggest that MSC apoptosis is crucial. They observed in a mouse model of graft-versus-host disease that cytotoxic cells rendered the MSCs apoptotic shortly after transfer. Moreover, cells from patients that responded to MSC therapy had more cytotoxic activity against MSCs. These findings not only provide important mechanistic insight but also suggest that patients could be screened for responsiveness to MSC therapy before transfer. The immunosuppressive activity of mesenchymal stromal cells (MSCs) is well documented. However, the therapeutic benefit is completely unpredictable, thus raising concerns about MSC efficacy. One of the affecting factors is the unresolved conundrum that, despite being immunosuppressive, MSCs are undetectable after administration. Therefore, understanding the fate of infused MSCs could help predict clinical responses. Using a murine model of graft-versus-host disease (GvHD), we demonstrate that MSCs are actively induced to undergo perforin-dependent apoptosis by recipient cytotoxic cells and that this process is essential to initiate MSC-induced immunosuppression. When examining patients with GvHD who received MSCs, we found a striking parallel, whereby only those with high cytotoxic activity against MSCs responded to MSC infusion, whereas those with low activity did not. The need for recipient cytotoxic cell activity could be replaced by the infusion of apoptotic MSCs generated ex vivo. After infusion, recipient phagocytes engulf apoptotic MSCs and produce indoleamine 2,3-dioxygenase, which is ultimately necessary for effecting immunosuppression. Therefore, we propose the innovative concept that patients should be stratified for MSC treatment according to their ability to kill MSCs or that all patients could be treated with ex vivo apoptotic MSCs.