Frequency Of Myeloid-derived Suppressor Cells Research Articles

Myeloid Derived Suppressor Cells Expansion Persists After Early ART and May Affect CD4 T Cell Recovery

Myeloid-derived suppressor cells (MDSC) are expanded during HIV-1 infection and correlated with disease progression. MDSC expand in the early phase of primary infection depending on TRAIL level. In this study we evaluated the effect of ART on the frequency of MDSC in patients with primary HIV infection (PHI), and their impact on CD4 T cell reconstitution. MDSC frequency was evaluated by flow-cytometry in 60 PHI patients at 12, 24 and 48 weeks after ART initiation. Cytokine plasma levels were evaluated by Luminex technology at the same time points. The capacity of MDSC to modulate hematopoietic early progenitor cells' expansion was evaluated using the OP9/Dl1 in vitro system. As previously described, polymorphonuclear-MDSC (PMN-MDSC) frequency was higher in PHI compared to healthy donors. Interestingly, 48 weeks of successful ART failed to normalize the PMN-MDSC frequency. Moreover, PMN-MDSC frequency was not correlated with residual viral load, suggesting that the persistence of PMN-MDSC was not due to residual viral replication. Interestingly, patients with low PMN-MDSC frequency (<6%) at T0 had a higher HIV DNA at the same time point than individuals with high PMN-MDSC frequency (>6%). We also found an inverse correlation between PMN-MDSC frequency and CD4-T cell count at 48 weeks post-ART, which was confirmed by multivariate analysis adjusting for age and CD4 T cell number at baseline. These data suggest that the persistence of PMN-MDSC may impact CD4 T cell recovery. Indeed, in vitro PMN-MDSC impaired the expansion of CD34+CD38- hematopoietic early progenitors. Further, a balance between TRAIL and GM-CSF may be necessary to maintain a low MDSC level. In conclusion, early ART initiation was not able to normalize PMN-MDSC frequency that might impact the CD4 T cell recovery. These data open new questions regarding the clinical impact of MDSC persistence in HIV+ patients, in particular on non-AIDS related diseases.

Abstract 4049: Cellular phenotypes of spleen cells in cancer patients targeted for adoptive cellular therapy

Abstract The spleen is a rich resource of large numbers of lymphocytes, with potential for use in adoptive cellular therapy (ACT). This is of particular relevance to patients with a distal pancreatic ductal adenocarcinoma (PDAC) who frequently undergo a pancreatectomy that required a splenectomy to clear the nodal basin. In this study we compared the phenotypes of eight paired sets of peripheral blood (PB) and spleen cell populations to assess the frequency of myeloid derived suppressor cells (MDSC) and lymphocyte subpopulations. Because a positive clinical response to ACT requires a low frequency of T-regs and MDSC’s, both systemically and in the tumor microenvironment these cells were one of the foci for our studies. A high frequency of these cells negatively correlate with survival and positively with tumor burden. Indeed, in this study we observed a significant decrease in the frequency of granulocytic, monocytic and immature MDSC’s in the spleen vs PB (4.8±1.6 vs. 54.4±6.1; 0.2±0.1 vs. 0.9±0.4; and 0.6±0.1 vs. 2.3±0.8 percent respectively) but not T-regulatory (T-reg) cells (0.8+0.5 vs. 0.1+0.04 percent) supporting the potential superiority of spleen cells relative to PB as a cellular source for ACT. In addition to a significantly higher number of spleen cells as compared to the PB for expansion/infusion, spleens have a higher frequency of T-cells as compared to the PB including CD3+ (35+5 vs. 16+4%), CD4+ (19+2 vs. 10+2%) and CD8+ (10+1 vs. 5+2%) T-cells. The frequency of transitional memory (Ttm) T-cells, which have a high proliferative potential were also significantly increased in the spleen vs. PB (37.5±6 vs. 26±3.0%) as was the frequency of PD1+CD8+ T-cells (8+2 vs 1+0.5%), which have been shown to have tumor specific cytotoxicity in cancer patients. In summary the increased frequency of PD-1+CD8+ cells, a Ttm memory phenotype and low levels of an exhaustion phenotype (TIM3 and Lag3 expression) in spleen cells vs the PB is supportive of the potential utility of these cells for expansion and therapeutic utility for these cells. Citation Format: Kathryn Cole, Quan Ly, Jesse L. Cox, Michael A. Hollingsworth, James C. Padussis, Jason M. Foster, Luciano M. Vargas, James E. Talmadge. Cellular phenotypes of spleen cells in cancer patients targeted for adoptive cellular therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4049.

Expansion of PMN-myeloid derived suppressor cells and their clinical relevance in patients with oral squamous cell carcinoma

ObjectivesOral squamous cell carcinoma (OSCC) is the most common head and neck malignancy worldwide, with a high mortality. The prognosis of OSCC remains unsatisfactory; the dysregulated immune system plays an important role in the pathogenesis of OSCC. Myeloid-derived suppressor cells (MDSCs) have been identified as immune-suppressive cells in multiple tumor types. The aim of this study was to clarify the underlying immunoregulatory mechanism of MDSC in patients with OSCC. Materials and methodsFlow cytometry was used to analyze the phenotype of MDSC among peripheral blood mononuclear cells (PBMCs) from patients with OSCC and healthy control subjects. The correlation between MDSC frequency and the disease index of patients with OSCC was evaluated. T cell proliferation experiment was used to evaluate the immunosuppressive function of MDSC. ResultsPatients with OSCC exhibited significantly higher levels of PMN-MDSCs than did healthy controls. In the co-culture assay, T cell proliferation and IFN-γ production were abrogated by the addition of PMN-MDSCs in a dose-dependent manner. The levels of reactive oxygen species were higher for PMN-MDSCs derived from patients with OSCC than for those from normal individuals. p-STAT3 levels, a key activator of MDSCs, was higher in OSCC-related PMN-MDSCs than in those from healthy controls. Both of these effects were reversed by NAC (an ROS inhibitor) and JSI-124 (a p-STAT3 inhibitor). Finally, PMN-MDSC levels were positively related to histological differentiation, nodal metastasis, and recurrence. ConclusionPMN-MDSCs were elevated in OSCC patients, with strong immune-suppressive effects via p-STAT3/reactive oxygen species, providing a new direction for therapeutic strategies.

Myeloid-Derived Suppressor Cells in Lung Transplantation.

Myeloid-derived suppressor cells (MDSC) are a heterogeneous group of immune cells from the myeloid lineage. MDSCs expand in pathological situations, such as chronic infection, cancer, autoimmunity, and allograft rejection. As chronic lung allograft dysfunction (CLAD) limits long-term survival after lung transplantation (LTx), MDSCs may play a role in its pathophysiology. We assessed phenotype and frequency of MDSCs in peripheral blood from lung transplant recipients and its relationship to post-transplant complications and immunosuppression. Granulocytic (G)-MDSC were identified and quantified by flow cytometry of blood from 4 control subjects and 20 lung transplant patients (stable n = 6, infection n = 5; CLAD n = 9). G-MDSC functionality was assessed in vitro by their capability to block CD4 and CD8 T cell proliferation. More G-MDSC could be assessed using EDTA tubes compared to heparin tubes (p = 0.004). G-MDSC were increased in stable lung transplant recipients vs. non-transplant controls (52.1% vs. 9.4%; p = 0.0095). The infection or CLAD groups had lower G-MDSCs vs. stable recipients (28.2%p = 0.041 and 33.0%; p = 0.088, respectively), but were not different among CLAD phenotypes. G-MDSC tended to correlate with cyclosporine A and tacrolimus levels (r2 = 0.18; r2 = 0.17). CD4 and CD8 cells proliferation decreased by 50 and 80% if co-cultured with MDSCs (1:6 and 1:2 MDSC:T-cell ratio, respectively). In conclusion, circulating MDSCs are measurable, functional and have a G-MDSC phenotype in lung transplant patients. Their frequency is increased in stable patients, decreased during post-transplant complications, and related to level of immunosuppression. This study may pave the way for further investigations of MDSC in the context of lung transplantation.

Myeloid-derived suppressor cells expansion is closely associated with disease severity and progression in HBV-related acute-on-chronic liver failure.

Dysregulation of the host immune responses induced by host hepatitis B virus (HBV) interactions has been observed in acute-on-chronic liver failure (ACLF). Myeloid-derived suppressor cells (MDSCs), well known for their immunomodulatory properties, can suppress T-cell function by regulating the expression of CD3 ζ chain in cancer and autoimmune/infectious diseases while rarely have been studied in ACLF. In this study, MDSCs, CD4+ /CD8+ T cells, and CD3 ζ chain were analyzed by flow cytometry in peripheral blood mononuclear cells obtained from HBV-related ACLF patients, chronic hepatitis B (CHB) patients and healthy controls. ACLF patients were followed up for dynamic detection of MDSCs and observation of outcomes after treatment. Interestingly, peripheral CD14+ CD33+ CD11b+ HLA-DR-/low MDSCs from ACLF patients were significantly increased compared to those from CHB patients and healthy controls. CD4+ /CD8+ T cell frequency and CD3 ζ chain expression in T cells were decreased in ACLF patients compared to those of healthy controls and were negatively correlated with matched MDSC frequency. Meanwhile, the frequency of MDSCs was closely correlated with biochemical parameters that are relevant for liver injury rather than virological parameters. Moreover, a lower level of MDSCs was correlated with a better short-term prognosis (within 4 weeks but not at 8 weeks), and MDSCs remained high in ACLF patients whose conditions worsened within a 4-week follow-up period after treatment. These results suggest that MDSCs are closely involved in cell-mediated immunity in HBV-related ACLF and that peripheral MDSC expansion is closely associated with disease severity and progression in HBV-related ACLF, which may serve as a predictor of short-term prognosis.

α-Difluoromethylornithine suppresses inflammatory arthritis by impairing myeloid-derived suppressor cells

ObjectivesThe chemopreventive drug α-difluoromethylornithine (DFMO) has been shown to have an antinociceptive effect on mechanical allodynia in inflammatory arthritis by directly inhibiting ornithine decarboxylase (ODC) and decreasing polyamine production in inflammatory sites. However, little is known about the effect of DFMO on the immune system of inflammatory arthritis. Here, we investigated the effect of DFMO in a well-established collagen-induced arthritis (CIA) mouse model and explored its effect on the immune system. MethodsThe effect of DFMO on the frequency of myeloid-derived suppressor cells (MDSCs) in the spleens of CIA mice and their associations with disease severity, tissue inflammation and the levels of proinflammatory T-helper (Th) 17 cells in lymphoid tissues were investigated. The effects of DFMO on disease severity and Th17 cells were compared with those of antibody depletion of MDSCs. The arthritis severity was also evaluated by adoptive transfer of MDSCs derived from DFMO- or dH2O-treated mice. ResultsIn this study, we showed that both MDSCs and Th17 cells were significantly expanded in CIA mice. Treatment by DFMO at the onset of CIA suppressed the development of arthritis and decreased the frequency of MDSCs and Th17 cells. MDSC depletion by anti-Gr-1 mAb achieved a similar result, while combination treatment of both methods did not achieve a significant difference compared to either of the single treatments. In addition, the adoptive transfer of MDSCs derived from dH2O-treated mice with CIA restored the arthritis severity of CIA in mice treated with anti-Gr-1 mAb, while the transfer of MDSCs from DFMO-treated mice did not have such an effect. ConclusionsOur results identified DFMO as a potential therapeutic drug for the treatment of inflammatory arthritis.

Frequency and Implications of myeloid-derived suppressor cells and lymphocyte subsets in Egyptian patients with hepatitis C virus-related hepatocellular carcinoma.

Myeloid-derived suppressor cells (MDSCs) play a pivotal role in tumor immunity and induction of immune tolerance to a variety of antitumor effectors, including T lymphocytes. Herein, we tried to evaluate the frequency and clinical significance of MDSCs and different lymphocyte subsets in hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC). Four groups were enrolled; chronic HCV (CHC; n = 40), HCV-related liver cirrhosis (n = 40), HCV-related HCC (HCV-HCC; n = 75), and healthy control group (n = 20). The percentage of peripheral lymphocytes subsets and total MDSCs with their main two subsets; monocytic (M-MDSCs) and granulocytic (G-MDSCs) was evaluated by flow cytometry. The frequency of total MSDCs and M-MDSCs was significantly elevated in HCV-HCC especially patients with advanced stage HCC compared with those with early-stage HCC. The frequency of total MSDCs and M-MDSCs was positively correlated with ALT, AFP, and HCV viral load and negatively correlated with CD8+ T-cell frequency. CD4 + T cells were significantly decreased in HCV-HCC patients. The frequency of CD4 + T cells and CD8 + T cells was negatively correlated with AFP and AST, but not with albumin or HCV viral load. Taken together, our data suggest that MDSCs, M-MDSCs, and lymphocyte subsets are associated with the development and progression of HCV-related HCC.

Myeloid-derived suppressor cells in the patients with liver resection for hepatitis B virus-related hepatocellular carcinoma

Liver resection remains the popular treatment for hepatocellular carcinoma (HCC). The aim of this study was to explore the alteration of immune cells in HCC patients with liver resections. Nineteen patients were included and their peripheral blood samples were taken before and after liver resections for immune-cell analysis. The clinical characteristics showed that the median diameter of the resected tumors was 7.5 cm with a range from 1.4 to 16.5 cm. The analysis of immune cells showed that the percentage of CD4+ T-cells were not altered by liver resection, but the percentage of CD8+ T-cell was decreased from 31.7 ± 12.4% to 20.2 ± 10.4% at one week after liver resection (p = 0.006). For immunosuppressor cells, regulatory T-cells were not altered, but myeloid-derived suppressor cells (MDSC) were decreased from 7.75 ± 8.16% to 1.51 ± 1.32% at one month after liver resection (p = 0.022) in 10 of 19 patients with high frequency of MDSC. Furthermore, it was also found that MDSC population was linearly correlated to tumor volume. In conclusion, CD8+ T-cells and MDSC were altered by liver resection. The percentage of CD8+ T-cells was decreased by surgery, but the accumulation of MDSC was abrogated.

Changes in the frequency of myeloid-derived suppressor cells after transarterial chemoembolization with gelatin sponge microparticles for hepatocellular carcinoma

PurposeA series of clinical studies have established the safety and efficacy of transcatheter arterial chemoembolization (TACE) with gelatin sponge microparticles (GSMs) in treating hepatocellular carcinoma (HCC). HCC can lead to obvious necrosis inside tumors, especially larger ones, although it is unclear whether such necrotic tumor tissue can induce favorable immune reactions against the tumor. Myeloid-derived suppressor cells (MDSCs) have immunosuppressive functions and are currently considered a very important cell type affecting tumor immunity. This study observed changes in MDSC frequency in peripheral blood before and after GSM–TACE to evaluate the effect on the immune function of HCC patients. MethodsEight patients diagnosed with HCC underwent GSM–TACE treatment in the Hepatobiliary Interventional Department of Beijing Tsinghua Chang Gung Hospital, Beijing, China; we followed up with the patients over a period of 30 days post-surgery. We used flow cytometry (FCM) to quantify the frequency of MDSCs in peripheral blood before TACE, 10 days after surgery and 30 days after surgery. ResultsMDSC frequency after GSM–TACE had a significant downward trend. Pre-TACE, it was 30.73% ± 11.93%, decreasing to 18.60% ± 11.37% at 10 days after operation. This decrease was not statistically significant (P > 0.05). MDSC frequency was even lower 30 days after TACE (7.63% ± 7.32%) than at 10 days after TACE (P < 0.05), and there was a significant difference compared with pre-TACE (P < 0.001). We evaluated tumor response at 30 days after GSM–TACE according to the Modified Response Evaluation Criteria in Solid Tumors (mRECIST), and all eight patients showed partial response (PR). ConclusionOur results confirmed that GSM–TACE was beneficial for improving anti-tumor immunity in the treatment of HCC.

T-cell-Secreted TNFα Induces Emergency Myelopoiesis and Myeloid-Derived Suppressor Cell Differentiation in Cancer.

Hematopoiesis in patients with cancer is characterized by reduced production of red blood cells and an increase in myelopoiesis, which contributes to the immunosuppressive environment in cancer. Some tumors produce growth factors that directly stimulate myelopoiesis such as G-CSF or GM-CSF. However, for a majority of tumors that do not directly secrete hematopoietic growth factors, the mechanisms involved in the activation of myelopoiesis are poorly characterized. In this study, we document in different murine tumor models activated hematopoiesis with increased proliferation of long-term and short-term hematopoietic stem cells and myeloid progenitor cells. As a consequence, the frequency of myeloid-derived suppressor cells and its ratio to CD8+ T cells increased in tumor-bearing mice. Activation of hematopoiesis and myeloid differentiation in tumor-bearing mice was induced by TNFα, which was mainly secreted by activated CD4+ T cells. Therefore, the activated adaptive immune system in cancer induces emergency myelopoiesis and immunosuppression. SIGNIFICANCE: These findings characterize a regulatory circuit linking activated T cells to suppression of tumor-specific immune responses, providing a conceptual advance in the understanding of emergency-hematopoiesis in cancer and opening new targets for therapeutic approaches. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/79/2/346/F1.large.jpg.

A Phase 2 Study to Assess the Immunomodulatory Capacity of a Lecithin-based Delivery System of Curcumin in Endometrial Cancer.

Curcumin is a botanical with anti-tumor and immunomodulatory properties. We hypothesized that curcumin supplementation might influence inflammatory biomarker levels in endometrial carcinoma (EC). In this open-label, non-randomized phase 2 study (NCT02017353), seven EC patients consumed 2 g/day Curcumin Phytosome (CP) orally for 2 weeks. Blood was taken at baseline, days 1, 7, 14, and 21. The following analytes were measured: curcuminoids and metabolites, 56 inflammatory biomarkers, COX-2, frequencies of myeloid-derived suppressor cells, dendritic cells and NK cells, expression of MHC molecules on leukocytes and monocytes and activation/memory status of T cells. Patients completed quality of life (QoL) questionnaires at baseline and end of treatment. Curcumin metabolites were detectable in plasma upon CP intake. CP downregulated MHC expression levels on leukocytes (P = 0.0313), the frequency of monocytes (P = 0.0114) and ICOS expression by CD8+ T cells (P = 0.0002). However, CP upregulated CD69 levels on CD16− NK cells (P = 0.0313). No differences were observed regarding inflammatory biomarkers, frequencies of other immune cell types, T cell activation and COX-2 expression. A non-significant trend to improved QoL was observed. Overall, CP-induced immunomodulatory effects in EC were modest without significant QoL changes. Given the small population and the observed variability in inter-patient biomarker levels, more research is necessary to explore whether benefits of CP can be obtained in EC by different supplementation regimens.Clinical Trial Registration: www.ClinicalTrials.gov, identifier NCT02017353; www.clinicaltrialsregister.eu, identifier 2013-001737-40.

Visualization and quantification of in vivo homing kinetics of myeloid-derived suppressor cells in primary and metastatic cancer.

Myeloid-derived suppressor cells (MDSCs) are immunosuppressive cells of the myeloid compartment and major players in the tumor microenvironment (TME). With increasing numbers of studies describing MDSC involvement in cancer immune escape, cancer metastasis and the dampening of immunotherapy responses, MDSCs are of high interest in current cancer therapy research. Although heavily investigated in the last decades, the in vivo migration dynamics of MDSC subpopulations in tumor- or metastases-bearing mice have not yet been studied extensively. Therefore, we have modified our previously reported intracellular cell labeling method and applied it to in vitro generated MDSCs for the quantitative in vivo monitoring of MDSC migration in primary and metastatic cancer. MDSC migration to primary cancers was further correlated to the frequency of endogenous MDSCs. Methods: Utilizing a 64Cu-labeled 1,4,7-triazacyclononane-triacetic acid (NOTA)-modified CD11b-specific monoclonal antibody (mAb) (clone M1/70), we were able to label in vitro generated polymorphonuclear (PMN-) and monocytic (M-) MDSCs for positron emission tomography (PET) imaging. Radiolabeled PMN- and M-MDSCs ([64Cu]PMN-MDSCs and [64Cu]M-MDSCs, respectively) were then adoptively transferred into primary and metastatic MMTV-PyMT-derived (PyMT-) breast cancer- and B16F10 melanoma-bearing experimental animals, and static PET and anatomical magnetic resonance (MR) images were acquired 3, 24 and 48 h post cell injection. Results: The internalization of the [64Cu]NOTA-mAb-CD11b-complex was completed within 3 h, providing moderately stable radiolabeling with little detrimental effect on cell viability and function as determined by Annexin-V staining and T cell suppression in flow cytometric assays. Further, we could non-invasively and quantitatively monitor the migration and tumor homing of both [64Cu]NOTA-αCD11b-mAb-labeled PMN- and M-MDSCs in mouse models of primary and metastatic breast cancer and melanoma by PET. We were able to visualize and quantify an increased migration of adoptively transferred [64Cu]M-MDSCs than [64Cu]PMN-MDSCs to primary breast cancer lesions. The frequency of endogenous MDSCs in the PyMT breast cancer and B16F10 melanoma model correlated to the uptake values of adoptively transferred MDSCs with higher frequencies of PMN- and M-MDSCs in the more aggressive B16F10 melanoma tumors. Moreover, aggressively growing melanomas and melanoma-metastatic lesions recruited higher percentages of both [64Cu]PMN- and [64Cu]M-MDSCs than primary and metastatic breast cancer lesions as early as 24 h post adoptive MDSC transfer, indicating an overall stronger recruitment of cancer-promoting immunosuppressive MDSCs. Conclusion: Targeting of the cell surface integrin CD11b with a radioactive mAb is feasible for labeling of murine MDSCs for PET imaging. Fast internalization of the [64Cu]NOTA-αCD11b-mAb provides presumably enhanced stability while cell viability and functionality was not significantly affected. Moreover, utilization of the CD11b-specific mAb allows for straightforward adaptation of the labeling approach for in vivo molecular imaging of other myeloid cells of interest in cancer therapy, including monocytes, macrophages or neutrophils.

Neutrophil and Granulocytic Myeloid-Derived Suppressor Cell-Mediated T Cell Suppression Significantly Contributes to Immune Dysregulation in Common Variable Immunodeficiency Disorders.

Common variable immunodeficiency disorders (CVID) represent a group of primary immunodeficiency diseases characterized by hypogammaglobulinemia and impaired specific Ab response, resulting in recurrent infections due to dysfunctional immune response. The specific mechanisms mediating immune deficiency in CVID remain to be determined. Previous studies indicated that immune dysregulation in CVID patients is associated with chronic microbial translocation, systemic immune activation, and altered homeostasis of lymphocytic and myeloid lineages. A detailed phenotypic, functional characterization of plasma markers and immune cell populations was performed in 46 CVID patients and 44 healthy donors. CVID patients displayed significantly elevated plasma levels of a marker of neutrophil activation neutrophil gelatinase-associated lipocalin. Neutrophils from CVID patients exhibited elevated surface levels of CD11b and PD-L1 and decreased levels of CD62L, CD16, and CD80, consistent with a phenotype of activated neutrophils with suppressive properties. Neutrophils from CVID patients actively suppressed T cell activation and release of IFN-γ via the production of reactive oxygen species. Furthermore, CVID was associated with an increased frequency of low-density neutrophils (LDNs)/granulocytic myeloid-derived suppressor cells. LDN/granulocytic myeloid-derived suppressor cell frequency in CVID patients correlated with reduced T cell responsiveness. Exogenous stimulation of whole blood with bacterial LPS emulated some but not all of the phenotypic changes observed on neutrophils from CVID patients and induced neutrophil population with LDN phenotype. The presented data demonstrate that neutrophils in the blood of CVID patients acquire an activated phenotype and exert potent T cell suppressive activity. Specific targeting of myeloid cell-derived suppressor activity represents a novel potential therapeutic strategy for CVID.

Pleurotus ferulae polysaccharides improve the antitumor efficacy of therapeutic human papillomavirus dendritic cell-based vaccine

ABSTRACTWe previously found that Pleurotus ferulae polysaccharides (PFPS) improved the maturation and function of dendritic cells (DCs). In this study, we investigated the effects of PFPS on the antitumor efficacy of therapeutic human papillomavirus (HPV) DC-based vaccine. PFPS stimulated DCs pulsed with HPV E6/E7 peptides were used to treat tumor mice on day 5 & 12 (HPV + PFPS-DCs early) and day 12 & 19 (HPV + PFPS-DCs late) after TC-1 cell injection. Compared to control group, both HPV + PFPS-DCs early and HPV + PFPS-DCs late strategies significantly inhibited tumor growth, which was significantly correlated with the increased activation status of both CD4+ and CD8+ T cells, the decreased frequencies of myeloid-derived suppressor cells, and the induction of HPV-specific CD8+ T cell responses. The survival of tumor mice was also greatly improved by HPV + PFPS-DCs early. Moreover, HPV + PFPS-DCs early completely inhibited the growth of second challenged TC-1 cells in tumor free mice. The results showed that PFPS improved the antitumor efficacy of therapeutic HPV DC-based vaccine, suggesting that PFPS might be a potential adjuvant for DC-based vaccines. This study provides a potential strategy for developing the therapeutic DC-based vaccine against cervical cancer.

Myeloid-derived suppressor cells and tumor: Current knowledge and future perspectives.

The immunosuppressive features of tumor lesions participate not only as one of the major players inducing cancer progression but also a big challenge for effective immunotherapy. It has been found that immunosuppression associated with chronic inflammatory factors, such as growth factors, cytokines, and chemokines is generated by stroma and tumor cells. Chronic and exhaustive secretion of these mediators triggers the generation of myeloid-derived suppressor cells (MDSCs) demonstrating one of the key players engaged in tumor immunosuppression. In point of fact, direct cell-to-cell contact is a prerequisite for immunosuppressive functions of MDSCs. From the clinical perspective, the frequency of peripheral blood MDSCs is correlated with clinical stage and therapeutic response in various cancers. Furthermore, MDSCs are involved in chemoresistant settings. Altogether, it is a rational therapeutic approach to block the fierce cycle in which MDSCs are developed and infiltrated to favor cancer progression. In this review, we will summarize recent findings of MDSCs in tumor progression and discuss potential therapeutic strategies that could be evaluated in future clinical trials.

Bendamustine with Total Body Irradiation Limits Murine Graft-Versus-Host Disease in Part Via Myeloid-Derived Suppressor Cells

BackgroundGvHD remains a significant challenge in allogeneic hematopoietic cell transplantation (HCT). An under-investigated strategy to reduce GvHD is the modification of the preparative conditioning regimen. Cyclophosphamide (CY) and total body irradiation (TBI) is a commonly utilized myeloablative conditioning regimen, but is associated with GvHD. Bendamustine (BEN) is an alkylating agent that has been applied recently in nonmyeloablative chemotherapy-based conditioning regimens, demonstrating reduced myelosuppression, transplant related mortality, and acute graft versus host disease (GvHD). We have recently published that BEN can effectively replace CY when given following murine haploidentical bone marrow transplantation. We therefore hypothesized that BEN can also have advantages over CY when given in conjunction with TBI as a myeloablative preparative regimen.MethodsWe used a major histocompatibility mismatched murine BMT model to evaluate GvHD following BEN-TBI and CY-TBI conditioning. BALB/c (H-2d) mice received equivalent doses of BEN or CY (based on maximum tolerated dose), followed by TBI (400 cGy) and infusion of bone marrow (BM) cells with splenocytes (SC) or purified T-cells from C57BL/6 (H-2b) mice. GvHD morbidity and mortality and engraftment were monitored and MDSC frequencies were assessed by flow cytometry and immunofluorescence.ResultsWe demonstrate that BEN-TBI conditioning, while facilitating complete donor chimerism, results in significantly less GvHD and improved survival compared to CY-TBI. In BEN-TBI conditioned mice, suppressive CD11b+Gr-1high myeloid cells are increased in the blood, bone marrow (BM), and spleen, with no evident differences in suppressive function. The ratio of Gr-1+ cells to T-cells is significantly increased in BEN-TBI mice in the intestines. When Gr-1high cells are depleted prior to transplant, the beneficial effects of BEN-TBI are partially lost. Alternatively, administration of G-CSF, which promotes CD11b+Gr-1+ myeloid cell expansion, trends towards an increase in survival in BEN-TBI mice.ConclusionsThese findings indicate a potential role of granulocytic myeloid-derived suppressor cells (MDSCs) in the mechanism by which BEN allows engraftment with reduced GvHD. BEN-TBI conditioning reduces GvHD morbidity and mortality compared to CY-TBI and may present a safer alternative to CY-TBI as a myeloablative conditioning regimen in allogeneic HCT. DisclosuresNo relevant conflicts of interest to declare.

Circulating Myeloid-Derived Suppressor Cells Reflect Mycosis Fungoides/Sezary Syndrome Disease Stage and Response to Treatment

Background: Despite mycosis fungoides (MF) and Sezary syndrome (SS) comprising the most common forms of cutaneous T cell lymphoma, the pathophysiology underlying these disorders remains poorly understood. Consequently, current prognostic guidelines based on disease spread exhibit wide variations in clinical outcome within each stage, underscoring an urgent need for novel approaches to MF/SS disease evaluation. A growing body of research suggests that systemic immune dysregulation represents an early, cardinal feature of MF/SS. We hypothesized that tracking this immune dysfunction in conjunction with disease spread may generate important pathophysiologic and prognostic information for patients. We focused on myeloid-derived suppressor cells (MDSCs), a recently discovered population of immunosuppressive innate immune cells related to neutrophils and monocytes, because their expansion in numerous solid tumor settings have correlated reliably with poor patient outcomes. Whether MF/SS augments circulating MDSC abundance remains unexplored, prompting us to evaluate whether this could serve as a marker for disease progression and treatment response.Methods: We used multiparametric flow cytometry to analyze the frequency and immunophenotype of MDSCs from the peripheral blood of 15 healthy donors and 30 patients with MF/SS. Patients at varying stages of MF/SS disease progression and treatment were included in the study. We defined granulocytic MDSCs (G-MDSCs) as cells positive for CD15, CD11b, and the recently discovered surface marker LOX-1, and negative for CD14. Monocytic MDSCs (M-MDSCs) were defined as cells positive for CD14 and CD11b, negative for CD15, and low/negative for HLA-DR. Each patient sample also underwent flow cytometry evaluating for circulating neoplastic T cells. These results were correlated with each participant's other hematologic parameters and clinical information through manual chart review.Results: We found that healthy donors harbored no quantifiable circulating MDSCs of either monocytic or granulocytic lineage, a result in keeping with previous studies. In contrast, MF/SS patients exhibited robust, statistically significant increases in the frequencies of both G-MDSCs and, to a lesser extent, M-MDSCs. G-MDSCs exceeded 20% of all CD15-positive cells in some patients.When patients were stratified by MF/SS clinical stage, those with more advanced disease displayed significantly higher G-MDSC abundance than early-stage patients. G-MDSC frequency was positively correlated with circulating CD4+ CD26- T cell counts often used in evaluating Sezary syndrome (R2 = 0.498; p < 0.0001). However, patients with early, skin-restricted disease also showed statistically significant increases in circulating G-MDSCs compared to healthy controls. This suggested that G-MDSC expansion may serve as a sensitive, blood-based disease marker even in the absence of systemic involvement by neoplasia.Patients who underwent recent treatment exhibited variable G-MDSC counts in the peripheral blood that were lower than in similar untreated patients on average. Serial measurements for two patients enrolled in a clinical trial for dual phosphoinositide 3-kinase and histone deacetylase inhibition revealed that G-MDSC frequencies markedly decreased over the course of treatment, mirroring the decrements of aberrant T cells circulating in the blood.Conclusion: These findings provide clear evidence of G-MDSC expansion in the peripheral blood of MF/SS patients that begins in early/locally restricted disease, grows with disease progression, and responds to systemic therapy. Such immunometric assays may illuminate a novel source of staging and prognostic information and may permit less invasive disease monitoring than current methods require. DisclosuresNo relevant conflicts of interest to declare.

The Pleiotropic Immunosuppressive Role of Osteoclasts in Multiple Myeloma

Skeletal related events impact both quality of life and overall survival of multiple myeloma (MM) patients. The underlying cause of osteolytic bone disease is an imbalance between bone-resorbing osteoclasts and bone-forming osteoblasts. Bone cells are regulated by a complex milieu of bone marrow microenvironmental components including the immune system via the effect of cytokines, signaling molecules and regulatory proteins. For example, osteoclastogenesis is tightly regulated by T cells through signaling crosstalk between RANKL and IFN-γ (Takayanagi H et al., Nature 2000). However, the effect of osteoclasts (OCs) on the immune system is less well defined.Here we investigated the effect of osteoclasts on immune cells including T cells, Th17 cells, NK cells and myeloid-derived suppressor cells (MDSCs) in MM. To gain insight into the functional impact of OCs on the immune cells, each type of immune cell was isolated from peripheral blood mononuclear cells (PBMNCs) and examined further in co-cultures with OCs. Our preliminary data showed that the frequency of NK cells and MDSCs in PBMNCs derived from myeloma patients increased when co-cultured with autologous OCs. Activation of the inhibitory immune checkpoints suppresses T cell-mediated antitumor immunity. Up-regulation of several co-inhibitory molecules has also been shown in MM (Kwon M et al., J Immunol 2017). We, therefore, assessed the expression of various immune checkpoint receptors, such as PD-1, LAG-3 and B7-H3, on T cells in co-culture with or without OCs by multi-color flow cytometry. OC co-culture significantly increased PD-1 expression in both CD4+and CD8+ T cell populations. The presence of OCs also enhanced PD-L1 protein expression on MM cells. Th17 cells, a newsubset of helper T cells, have recently been identified as immunosuppressive cells. Th17 cells are increased in myeloma; morevover they secrete IL-17 which promotes myeloma cell growth (Prabhala RH et al., Blood 2010). Interestingly, we found that naïve CD4 T cells have a higher propensity to differentiate into Th17 lineage in the presence of Th17 differentiation cytokines when co-cultured with OCs. Furthermore, we also observed an increased expansion of Th17 cells in co-culture with OCs. NK cell cytotoxic function is severely impaired in myeloma (Fauriat C et al., Leukemia. 2006). In our experiments, NK cells co-cultured in the presence of OCs demonstrated significant decrease in NK cell cytotoxicity, despite increase in NK cell numbers.Our study demonstrated that OCs suppress the cytotoxic function of T cells and NK cells and induce the expansion of immunosuppressive Th17 cells and MDSCs through direct contact. These findings indicate that OCs play an important role in tumor progression in part by enhancing immune suppression. Targeting OCs represent an attractive therapeutic option not only for controlling osteolytic bone disease but also for restoring the impaired immune surveillance in MM. Ongoing studies will focus on understanding direct-contact-mediated interactions between OCs and immune cells. Furthermore, the effect of OC inhibition using anti-resorptive drugs on the immune system will be interrrogated. DisclosuresRaje:Medscape: Honoraria; Research to Practice: Honoraria; AstraZeneca: Research Funding; Takeda: Consultancy; Merck: Consultancy; Janssen: Consultancy; Celgene: Consultancy; BMS: Consultancy; Amgen Inc.: Consultancy.

Pre-Transplant Monocytic Myeloid-Derived Suppressor Cell Frequency Has No Prognostic Role for Outcome after Allogeneic Hematopoietic Cell Transplantation for Acute Myeloid Leukemia in Remission

INTRODUCTION: Emerging data indicate myeloid-derived suppressor cells (MDSCs) adversely impact outcomes of patients with a variety of malignancies. However, it is less clear how MDSCs affect acute myeloid leukemia (AML) disease biology or the response of AML patients to treatment. Here, we studied a cohort of people with AML undergoing hematopoietic cell transplantation (HCT) in first complete remission (CR1) to examine the relationship between the monocytic subtype of MDSCs (M-MDSCs), pre-transplant measurable residual disease (MRD) status, and outcome after HCT.PATIENTS AND METHODS: Adults ≥18 years of age were included if they underwent first allogeneic HCT with myeloablative or nonmyeloablative conditioning for AML in CR1 from April 2006 until October 2014. Prospective MRD testing in bone marrow aspirates was performed routinely via 10-color multiparameter flow cytometry (MFC) as part of the pre-transplant work-up. Pre-transplant MFC data were used retrospectively to quantify M-MDSCs as the proportion of monocytes (identified by side scatter properties and CD14 positivity) with low or negative expression of HLA-DR. M-MDSC frequency was then expressed as a percentage of the total number of viable cells in the sample. Available combinations of antigens measured by MFC did not allow for the identification of other MDSC subsets (e.g. granulocytic MDSCs).RESULTS: We identified 349 adults undergoing allogeneic HCT for AML in CR1 for whom pre-transplant MRD testing was performed and for whom follow-up data were available. Of these, 8 had to be excluded because of insufficient MFC events available for identification of M-MDSCs. In the remaining 341 patients, M-MDSC frequency ranged from <0.001% to 6.53%, with a median of 0.18% (25th percentile: 0.06%; 75th percentile: 0.40%). Patients with lower (i.e. <median) frequency of M-MDSCs were less likely male (p=0.01) and more likely had recovered neutrophil and platelet counts (p=0.001) than patients with higher (i.e. ≥median) frequency of M-MDSCs. There was no difference with regard to all other examined characteristics (age, WBC and cytogenetic risk at diagnosis, type of AML, median CR duration before HCT, donor type, and conditioning intensity). The proportion of MRD positive (“MRDpos”) patients was similar between those with lower vs. higher M-MDSC frequency (18.8 vs. 24.6%, p=0.20). Moreover, while MRD negative (“MRDneg”) patients (n=267) differed statistically significantly from MRDpos patients (n=74) with regard to several characteristics such as more favorable disease risk and lower proportion of secondary AML, there was only a statistically non-significant trend toward lower M-MDSC frequency in MRDneg patients (0.16% [range <0.001-5.61%] vs. 0.22% [range <0.001-6.53%], p=0.09).The median follow-up time after HCT among all survivors was 5.8 (range, 2.0-11.4) years. The time was slightly longer for survivors with lower compared to those with higher M-MDSC frequency (6.1 [range 2.6-11.4] vs. 5.1 [range 2.0-11.3] years; p=0.0076). Stratified by the median M-MDSC frequency, the 3-year estimates of overall survival were similar for patients with lower and higher pre-HCT M-MDSCs (58.2% [95% confidence interval 50.4-65.2%] vs. 57.6% [49.8-64.7%]), as were 3-year estimates for relapse free survival (53.5% [45.7-60.7%] vs. 48.3% [40.6-55.6%]), 3-year estimates of the cumulative incidence of relapse (31.8% [24.9-38.8%] vs. 34.6% [27.6-41.8%]), and 3-year estimates of non-relapse mortality (14.7% [9.9-20.5%] vs. 17.0% [11.8-23.0%]). Analyses restricted to the subset of patients undergoing myeloablative (MA) conditioning or those undergoing MA conditioning in MRDneg remission showed qualitatively similar results.CONCLUSIONS: In the largest study to date examining the role of M-MDSCs in AML, we found no convincing evidence for a prognostic role of these cells for adults undergoing allografting in CR1. DisclosuresWalter:Actinium Pharmaceuticals, Inc.: Other: Clinical trial support, Research Funding; Amgen Inc.: Other: Clinical trial support, Research Funding; Amphivena Therapeutics: Consultancy, Equity Ownership, Other: Clinical trial support, Research Funding; Aptevo Therapeutic: Consultancy, Other: Clinical trial support, Research Funding; Covagen AG: Consultancy, Other: Clinical trial support, Research Funding; Seattle Genetics, Inc.: Consultancy, Other: Clinical trial support, Research Funding; Boehringer Ingelheim Pharma GmbH & Co. KG: Consultancy; Pfizer: Consultancy.

IMMU-70. GLOBAL IMMUNE FINGERPRINTING IN GLIOBLASTOMA REVEALS IMMUNE-SUPPRESSION SIGNATURES ASSOCIATED WITH PROGNOSIS

Glioblastoma (GBM) remains uniformly lethal, and, despite a large accumulation of immune cells in the microenvironment, there is limited anti-tumor immune response, even with newly developed immune checkpoint therapies. To overcome these challenges and enhance the efficacy of immunotherapies, a comprehensive understanding of the immune system in GBM and changes during disease progression is required. Here, we integrated multi-parameter flow cytometry and mass cytometry time of flight (CyTOF) analysis of patient blood to determine changes in the immune system among tumor types and over disease progression. Utilizing multi-parameter flow cytometry analysis in a cohort of over 250 patients with brain tumors ranging from benign to malignant primary and metastatic, we found that GBM patients had a significant elevation in myeloid-derived suppressor cells (MDSCs) in blood, but not immunosuppressive T regulatory cells. We validated these findings in GBM patient tissue and found that increased numbers of MDSCs in recurrent GBM portended poor prognosis. CyTOF analysis of peripheral blood from a cohort of newly diagnosed GBM patients revealed that reduction in MDSC frequency over time is accompanied by a concomitant increase in dendritic cells and natural killer cells. This reduced MDSC profile was present in GBM patients with extended survival and was similar to that of low-grade glioma (LGG) patients. Our findings provide a rationale for developing strategies to target MDSCs, which are elevated in GBM patients and predict poor prognosis, either by directly targeting or by shifting the immune profile to induce differentiation toward the immune profile of LGGs.