Identification of a new subset of myeloid suppressor cells in peripheral blood of melanoma patients and modulation by GM-CSF-based anti-tumor vaccine

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 21082-21082
Author(s):  
P. Filipazzi ◽  
R. Valenti ◽  
V. Huber ◽  
M. Iero ◽  
L. Pilla ◽  
...  
Keyword(s):  
T Cell ◽  
Peripheral Blood ◽  
Tumor Vaccine ◽  
Suppressor Cells ◽  
Suppressive Activity ◽  
Gm Csf ◽  
Myeloid Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors ◽  
Melanoma Patients

21082 Background: Phenotypic and functional features of myeloid suppressor cells (MSC), known to serve as critical regulators of anti-tumor T cell responses in tumor-bearing mice, are still poorly defined in human cancers. Here we analyzed myeloid subsets with suppressive activity present in peripheral blood of metastatic melanoma patients (MM) and evaluated their modulation by a GM-CSF-based anti- tumor vaccine. Methods: Stage IV AJCC MM patients (n=16) vaccinated with autologous tumor-derived heat-shock protein peptide complexes gp96 (HSPPC-96) and low dose GM-CSF provided pre- and post-treatment whole blood samples. Peripheral blood mononuclear cells (PBMC) were analyzed by flow cytometry, separated into cellular subsets and used for in vitro proliferation assays. PBMC from stage- matched melanoma patients (n=12) treated with non-GM-CSF-based vaccines (i.e. HSPPC-96 alone or IFNa/melanoma-derived peptides) or gender and age-matched healthy donors (n=16) were also analyzed for comparison. Results: The lack or low levels of HLA-DR expression was found to identify a CD14+ cell subset with high suppressive activity on lymphocyte proliferation and functions. CD14+HLA-DR-/lo cells were significantly expanded in all MM patients, while undetectable in healthy donors. Suppressive activity was mediated by TGFβ, as suggested by functional experiments with neutralizing specific antibodies. In contrast, no involvement of arginase and iNOS pathways could be detected. CD14+HLA-DR-/lo cells, as well as spontaneous ex- vivo release and plasma levels of TGFβ, were augmented after administration of the HSPPC-96/GM-CSF vaccine. Interestingly, the expansion of suppressive CD14+ monocytes was associated to the inability to mount a significant CD8-mediated T cell response upon vaccination. On the other hand, no quantitative or qualitative enhancement of the CD14+HLA-DR-/lo suppressive cell population was observed in patients receiving a non-GM-CSF based vaccine. Conclusions: CD14+HLA-DR- /lo cells exerting TGFβ-mediated immune suppression may represent a new subset of myeloid suppressive cells, potentially expandable by the administration of GM-CSF-based vaccines in melanoma patients. [Table: see text]

Identification of a New Subset of Myeloid Suppressor Cells in Peripheral Blood of Melanoma Patients With Modulation by a Granulocyte-Macrophage Colony-Stimulation Factor–Based Antitumor Vaccine

2007 ◽  
Vol 25 (18) ◽  
pp. 2546-2553 ◽  
Author(s):  
Paola Filipazzi ◽  
Roberta Valenti ◽  
Veronica Huber ◽  
Lorenzo Pilla ◽  
Paola Canese ◽  
...  
Keyword(s):  
Metastatic Melanoma ◽  
Peripheral Blood ◽  
Suppressor Cells ◽  
Suppressive Activity ◽  
Gm Csf ◽  
Myeloid Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors ◽  
Melanoma Patients ◽  
Macrophage Colony

Purpose Phenotypic and functional features of myeloid suppressor cells (MSC), which are known to serve as critical regulators of antitumor T-cell responses in tumor-bearing mice, are still poorly defined in human cancers. Here, we analyzed myeloid subsets with suppressive activity present in peripheral blood of metastatic melanoma patients and evaluated their modulation by a granulocyte-macrophage colony-stimulating factor (GM-CSF) –based antitumor vaccine. Patients and Methods Stage IV metastatic melanoma patients (n = 16) vaccinated with autologous tumor-derived heat shock protein peptide complex gp96 (HSPPC-96) and low-dose GM-CSF provided pre- and post-treatment whole blood specimens. Peripheral-blood mononuclear cells (PBMCs) were analyzed by flow cytometry, separated into cellular subsets, and used for in vitro proliferation assays. PBMCs from stage-matched metastatic melanoma patients (n = 12) treated with non–GM-CSF-based vaccines (ie, HSPPC-96 alone or interferon alfa/melanoma–derived peptides) or sex- and age-matched healthy donors (n = 16) were also analyzed for comparison. Results The lack of or low HLA-DR expression was found to identify a CD14+ cell subset highly suppressive of lymphocyte functions. CD14+HLA-DR–/lo cells were significantly expanded in all metastatic melanoma patients, whereas they were undetectable in healthy donors. Suppressive activity was mediated by transforming growth factor beta (TGF-β), whereas no involvement of the arginase and inducible nitric oxide synthase pathways could be detected. CD14+HLA-DR–/lo cells, as well as spontaneous ex vivo release and plasma levels of TGF-β, were augmented after administration of the HSPPC-96/GM-CSF vaccine. No enhancement of the CD14+-mediated suppressive activity was found in patients receiving non–GM-CSF-based vaccines. Conclusion CD14+HLA-DR–/lo cells exerting TGF-β–mediated immune suppression represent a new subset of MSC potentially expandable by the administration of GM-CSF–based vaccines in metastatic melanoma patients.

Myeloid Impairment Contributes to Immunoparesis in Multiple Myeloma but Not MGUS

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1831-1831
Author(s):  
Alessandra Romano ◽  
Nunziatina Parrinello ◽  
Calogero Vetro ◽  
Piera La Cava ◽  
Annalisa Chiarenza ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
T Cell ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Healthy Subjects ◽  
Conflicts Of Interest ◽  
Myeloid Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors

Abstract Abstract 1831 Introduction In Multiple Myeloma (MM), but not in the monoclonal gammopathy of unknown significance (MGUS), the immune function is impaired as consequence of an immunologically hostile microenvironment and cellular defects, including reduction of immuno-surveillance and T-cell immunoparesis. We conducted an study focused on the myeloid compartment in MM, and its role in the progression from MGUS to MM. Methods Between January 2009 and April 2011 peripheral blood obtained from 60 consecutive newly diagnosed MM and 70 MGUS plus 30 healthy subjects was studied for evaluation of myeloid subpopulations and lymphoid paresis. Myeloid dysfunction was evaluated as percentage and absolute count of circulating myeloid suppressor cells (MDSC) in peripheral blood assessed by flow cytometry as follows: im-MDSC (CD34+/CD11b+/CD13+/CD14-/ HLA-DR-/CD45+), neutrophilic-like N-MDSC (CD11b+/CD13+/CD15+/CD14-/HLA-DR-/Lin-) and monocytic-like mo-MDSC (CD14+/HLA-DRlow/-). Myeloid function was evaluated by phagocytic activity using a commercially available kit (Phagotest R). Further, we investigated whether MM-neutrophils were able to induce anergy in T-cells. Neutrophils isolated from healthy donor (N=6), MGUS (N=3) or MM (N=6) peripheral blood (PB) were co-cultured with T-lymphocytes obtained from healthy donors. Expression of markers of activation in response to stimulation with PHA-P for 2 hours was assessed by flow cytometry as antigen density expressed as normalized mean of fluorescence intensity (N-MFI) of CD71 at 48 hours. Results The capability of phagocytosis of in neutrophils and monocytes from MM patients at diagnosis was significantly reduced compared to healthy subjects (p<0.001) and MGUS (p<0.0001). While the mature suppressive N-MDSC subset was not increased in MGUS and MM patients, the mo-MDSC subpopulation showed an increasing trend from healthy donors through MM (p=0.06) and the im-MDSC subset was significantly higher in MM vs healthy (p=0.002) and MGUS (p=0.001). After PHA-P stimulation, expression of CD71 (a marker of activation) in normal T-lymphocytes was increased (2954 ± 240.6 arbitrary units, au), and it was reduced (751.3 ± 30.48 au, p=0.0001) when co-coltured with MM-neutrophils, while no differences were evident in co-colture with MGUS- (2783 ± 206.1 au, p=0.61) or healthy donors-neutrophils (2588 ± 135.4, p=0.38). Conclusion Taken together, our findings suggest that in MM but not in MGUS there is a myeloid cell dysfunction that is correlated to impairment of T- cell arm. These alterations may have a role in the development of MM. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Regulatory T Cells Fail to Suppress Fast Homeostatic Proliferation In Vitro

Life ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 245
Author(s):  
Daniil Shevyrev ◽  
Valeriy Tereshchenko ◽  
Elena Blinova ◽  
Nadezda Knauer ◽  
Ekaterina Pashkina ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Autoimmune Diseases ◽  
Peripheral Blood ◽  
Treg Cells ◽  
Homeostatic Proliferation ◽  
Suppressive Activity ◽  
Healthy Donors

Homeostatic proliferation (HP) is a physiological process that reconstitutes the T cell pool after lymphopenia involving Interleukin-7 and 15 (IL-7 and IL-15), which are the key cytokines regulating the process. However, there is no evidence that these cytokines influence the function of regulatory T cells (Tregs). Since lymphopenia often accompanies autoimmune diseases, we decided to study the functional activity of Tregs stimulated by HP cytokines from patients with rheumatoid arthritis as compared with that of those from healthy donors. Since T cell receptor (TCR) signal strength determines the intensity of HP, we imitated slow HP using IL-7 or IL-15 and fast HP using a combination of IL-7 or IL-15 with anti-CD3 antibodies, cultivating Treg cells with peripheral blood mononuclear cells (PBMCs) at a 1:1 ratio. We used peripheral blood from 14 patients with rheumatoid arthritis and 18 healthy volunteers. We also used anti-CD3 and anti-CD3 + IL-2 stimulation as controls. The suppressive activity of Treg cells was evaluated in each case by the inhibition of the proliferation of CD4+ and CD8+ cells. The phenotype and proliferation of purified CD3+CD4+CD25+CD127lo cells were assessed by flow cytometry. The suppressive activity of the total pool of Tregs did not differ between the rheumatoid arthritis and healthy donors; however, it significantly decreased in conditions close to fast HP when the influence of HP cytokines was accompanied by anti-CD3 stimulation. The Treg proliferation caused by HP cytokines was lower in the rheumatoid arthritis (RA) patients than in the healthy individuals. The revealed decrease in Treg suppressive activity could impact the TCR landscape during lymphopenia and lead to the proliferation of potentially self-reactive T cell clones that are able to receive relatively strong TCR signals. This may be another explanation as to why lymphopenia is associated with the development of autoimmune diseases. The revealed decrease in Treg proliferation under IL-7 and IL-15 exposure can lead to a delay in Treg pool reconstitution in patients with rheumatoid arthritis in the case of lymphopenia.

A Population of Suppressive Monocytes Inhibiting T Cell Proliferation and Dendritic Cell Differentiation in Relapsed Non-Hodgkin’s Lymphoma

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 808-808
Author(s):  
Yi Lin ◽  
Peggy A Bulur ◽  
Michael P. Gustafson ◽  
Thomas E. Witzig ◽  
Allan B. Dietz
Keyword(s):  
Cell Proliferation ◽  
T Cell ◽  
Peripheral Blood ◽  
Hodgkin’S Lymphoma ◽  
Hodgkin's Lymphoma ◽  
T Cell Proliferation ◽  
Hla Dr ◽  
Healthy Donors ◽  
Non Hodgkin's Lymphoma ◽  
Kinase Phosphorylation

Abstract Despite advances in treatments for patients with non-Hodgkin’s lymphoma (NHL), the relapse rates remain high and 40% of diffuse large B-cell NHL (DLBCL) patients die of disease. New therapies to augment the host anti-tumor immune response are needed. Reports of graft-versus-lymphoma responses in patients who have received allogeneic hematopoietic cell transplant indicate a role for cellular immunotherapy. However, these patients have variable levels of immunodeficiency which may impact the efficacy of cellular therapy. To study this we first evaluated the cellular immune status of patients with relapsed NHL. Proliferation of peripheral blood mononuclear cells (PBMNC) stimulated with anti-CD3/CD28 beads was reduced by more than 3.5 folds for patients with DLBCL (n = 3) compared to that of age-matched healthy donors (n = 5; p = 0.02). Removal of monocytes from PBMNC by use of anti-CD14 immunomagnetic beads restored proliferation to that of healthy donors. Further, monocytes from these patients were deficient in stimulating allogeneic T cell proliferation by 3 folds compared to monocytes from healthy donors (n = 3 NHL; n = 8 normal; p < 0.01). Peripheral blood from 12 NHL patients (9 DLBCL; 1 grade 3 follicular lymphoma; 2 composite) and 12 age-matched healthy donors were characterized by flow cytometry to determine the phenotype of these suppressive monocytes. There was no difference in the % monocytes in the blood between NHL patients and healthy donors; however, NHL patients had elevated % monocytes with a suppressive phenotype (CD14+HLA-DRneg) compared to normals (NHL 38.9 ± 4.93%; normal 8.3 ± 2.15; p < 0.0001). This phenotype is distinct from other myeloid suppressors (Lin-CD33+HLA-DR-) or non-classical monocytes (CD16+), neither of which was different in numbers between NHL and normal donors. This suggests that the CD14+HLA-DRneg monocytes are responsible for the observed T cell suppression. To further characterize the function of these cells, we cultured purified CD14+ monocytes from NHL and compared their differentiation capacity with those from normal donors. The percentage of CD14+HLA-DRneg monocytes in initial ex vivo culture was inversely correlated with the percentage of pure, mature dendritic cells (mDC) generated with TNF-a and PGE2 as maturation factors (CD80+CD83+; n = 9; p = 0.015). As CpG oligonucleotides are also capable of immune stimulation and have some anti-tumor activity in clinical trials, we investigated the effect of CpG on mDC differentiation in NHL. In healthy donors, maturation of monocytes with CpG yielded highly pure mDC (90.7 ± 2.15%, n = 3). However, preliminary results of mDC yield from monocytes of NHL patients matured with CpG was only 22.6 ± 11.2% (n = 2). This data suggests alternative signaling pathways of these suppressive monocytes. Preliminary analysis of a proteome array for 46 kinase phosphorylation sites from 37 proteins in 2 NHL and 3 healthy donors suggest changes in phosphorylation of 17 protein kinases for CD14+HLA-DRneg compared to CD14+HLA-DR+ cells. Functional correlation of these protein kinase phosphorylation changes is needed to definitively target the pathway characteristic of CD14+HLA-DRneg monocytes. Finally, we have identified a serum-free culture method that can consistently generate highly pure mDC from monocytes of NHL patients (93.0 ± 3.6%, n = 9) and is readily adaptable to good manufacturing practice for clinical use in immunotherapy. Taken together we have described for the first time a population of CD14+HLA-DRneg monocytes that is a significant source of immunosuppression in NHL patients and are beginning to target methods of overcoming this suppression.

scholarly journals Myeloidderived peripheral blood suppressor cells at haematopoietic stem cell mobilisation in multiple myeloma patients

2021 ◽  
Vol 66 (2) ◽  
pp. 218-230
Author(s):  
T. A. Aristova ◽  
E. V. Batorov ◽  
V. V. Sergeevicheva ◽  
S. A. Sizikova ◽  
G. Yu. Ushakova ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Stem Cell ◽  
Partial Response ◽  
Peripheral Blood ◽  
Plasma Cells ◽  
Suppressor Cells ◽  
Complete Response ◽  
Haematopoietic Stem Cell ◽  
Hla Dr ◽  
Healthy Donors

Introduction. Multiple myeloma (MM) is a B-cell malignancy with clonal expansion of plasma cells in bone marrow. Highdose chemotherapy with autologous haematopoietic stem cell transplantation is among main consolidation therapies in MM. Myeloid-derived suppressor cells (MDSCs) are immature myeloid-accompanying cells able to suppress the immune response. The administration of granulocyte colony stimulating factor (G-CSF) to mobilise haematopoietic stem cells (HSCs) increases the MDSC count in peripheral blood (PB).Aim — to study MDSC subsets in PB of remission MM patients and their incidence dynamics at HSC mobilisation.Methods. The study surveyed 35 MM patients prior to and after HSC mobilisation. The counts of granulocytic (G-MDSCs; Lin–HLA-DR–CD33+ CD66b+), monocytic (М-MDSCs; CD14+ HLA-DRlow/–) and early MDSCs (E-MDSCs; Lin–HLA-DR– CD33+ CD66b–) were estimated in flow cytometry.Results. Remission MM patients differed from healthy donors in higher relative counts of G-MDSCs (Lin–HLA-DR– CD33+ CD66b+) and increased relative and absolute counts of М-MDSCs (CD14+ HLA-DRlow/–). М-MDSCs significantly outnumbered G-MDSCs. MDSC subset counts were elevated in complete response (CR) and very good partial response (VGPR), as well as in partial response (PR). Higher relative MDSC counts were associated with greater pretreatment (2–3 lines of chemotherapy). After HSC mobilisation with cyclophosphamide 2–4 g/m2 + G-CSF (filgrastim 5 μg/kg/day), the median relative E-MDSC and M-MDSC counts increased by 2.3 and 2.0 times, respectively, while the relative G-MDSC count raised 46-fold perturbing the MDSC subset balance.Conclusion. Remission MM patients had the increased relative G-MDSC and both relative and absolute M-MDSC counts compared to donors. A greater patient pretreatment was associated with higher relative G-MDSC counts. Treatment response (CR/VGPR vs. PR) was not coupled with MDSC count variation. The G-CSF-induced HSC mobilisation entailed a significant expansion of all three MDSC subsets in PB.

scholarly journals Myeloid-Derived Suppressor Cells (HLA-DR-/lowCD16-CD33+) Expanded By Granulocyte Colony-Stimulating Factor Can Prevent Acute Graft-Versus-Host Disease in Humanized Mouse and Patients Following HSCT

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 72-72
Author(s):  
Ke Wang ◽  
Meng Lv ◽  
Ying-Jun Chang ◽  
Xiang-Yu Zhao ◽  
Xiao-Su Zhao ◽  
...  
Keyword(s):  
Graft Versus Host Disease ◽  
Peripheral Blood ◽  
Acute Gvhd ◽  
Suppressor Cells ◽  
Myeloid Derived Suppressor Cells ◽  
Granulocyte Colony Stimulating Factor ◽  
Graft Versus Host ◽  
Hla Dr ◽  
Healthy Donors

Abstract Introduction Myeloid-derived suppressor cells (MDSCs) are proposed to control graft-versus-host disease (GVHD) in allogeneic hematopoietic stem cell transplantation (allo-HSCT). However, the definition of human MDSCs has not yet reached consensus. Granulocyte colony-stimulating factor (G-CSF) has been routinely used to mobilize stem cells to peripheral blood in healthy donors. It was also recognized as a novel mediator of T-cell tolerance. However, the effects of G-CSF administration on donor-derived MDSCs and the further regulatory effects of these MDSCs on GVHD remained unclear. Amis The aim of this study is to evaluate the in vitro and in vivo effects of G-CSF expanded, donor-derived MDSCs (HLA-DR-/lowCD16-CD33+) in preventing acute GVHD after allo-HSCT. Methods The frequency and cell numbers of different kinds of MDSCs in peripheral blood before and after G-CSF administration from 10 healthy donors were analyzed by flow cytometry. Cells morphological features were detected by May-Grünwald-Giemsa cytospin. Secondly, the suppressive and regulatory functions of HLA-DR-/lowCD16-CD33+ population on CD3+ T cells were assessed via in vitro experiments. A humanized xenogeneic acute GVHD model was established to determine whether this population could prevent acute GVHD in vivo. Furthermore, a clinical prospective cohort study enrolled one hundred consecutive transplant recipients was performed to assess the effects of HLA-DR-/lowCD16-CD33+ contained in HSC grafts on the occurrence of acute GVHD. Results The findings of this study include: First, a novel phenotype of HLA-DR-/lowCD16-CD33+ MDSCs with suppressive function and morphological features similar to those of immature monocyte was identified. The median of percentages of this subset were significantly increased both in peripheral blood (PB, 6.5% vs. 4.6%, P=0.0122) and peripheral blood stem cells harvest (PBSCs, 15.5% vs. 4.6%, P<0.0001) after treating healthy donors with G-CSF than those of PB before mobilization. The median of percentage of interleukin-10 (IL-10) and transforming growth factor beta (TGF-β) positive cells among HLA-DR-/lowCD16-CD33+ population were both significantly higher than those of PB before mobilization. (IL-10+, 10.1% vs.0.62%, P=0.002; TGF-β+, 60.24% vs. 10.39%, P=0.0003). Donor-derived HLA-DR-/lowCD16-CD33+population inhibited the proliferation of T cells in a TGF-β-dependent manner. In vitro cell co-culture experiments also showed that this MDSCs subset could promote regulatory T cells (Treg, CD4+CD25+Foxp3+) expansion and induce T helper 2(Th2, CD4+IL-4+) differentiation (The median of percentage of Treg in co-culture with MDSCs group, 21.4% vs. without MSDCs group, 8.35%, P=0.0048. The median of fold change of Th2/Th1 in co-culture with MDSCs group, 1.35 vs. without MSDCs group 0.98, P=0.0159. The median of fold change of Th2/(Th1+Th17) in co-culture with MDSCs group, 1.28 vs. without MSDCs group 1.00, P=0.0095. Th1 (CD4+IFNγ+), Th17 (CD4+IL-17A+)). Second, we demonstrated that these cells could prevent acute GVHD in a humanized mouse model. Adoptive transfer human G-CSF-mobilized HLA-DR-/lowCD16-CD33+ cells significantly prolonged the survival and ameliorated the weight loss and tissue damage in GVHD mice. Third, clinical cohort results showed that the number of HLA-DR-/lowCD33+CD16- cells in the donor graft was the only independent risk factor inversely correlated with the incidence II-IV acute GVHD in recipients (HR 0.388, 95% CI: 0.158-0.954, P=0.039). Conclusion Our results suggest that MSDCs with HLA-DR-/lowCD16-CD33+ phenotype in G-CSF-mobilized PBSCs have monocytic features and immune-regulatory properties, which could alleviate acute GVHD in the allo-HSCT settings. Key words: Myeloid-derived suppressor cells; granulocyte colony-stimulating factor; graft-versus-host disease Figure Figure. Disclosures No relevant conflicts of interest to declare.

scholarly journals EFFECT OF HLA-DR ANTIBODY CONCENTRATION ON ESTIMATION OF THE NUMBER OF CD14+HLA-DRLOW /–-CELLS IN THE BLOOD OF PATIENTS WITH B-CELL chronic lymphocytic leukemia

2020 ◽  
Vol 19 (3) ◽  
pp. 65-67
Author(s):  
A. V. Ponomarev ◽  
V. A. Misyurin ◽  
M. A. Baryshnikova
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Peripheral Blood ◽  
Suppressor Cells ◽  
Lymphocytic Leukemia ◽  
Antibody Concentration ◽  
Myeloid Derived Suppressor Cells ◽  
Hla Dr ◽  
Healthy Donors ◽  
Cell Chronic Lymphocytic Leukemia

Introduction. Immunoregulatory functions of myeloid-derived suppressor cells have been extensively studied over the recent decades. Additionally, myeloid-derived suppressor cells have been investigated as a prognostic factor.Objective. To optimize the measurement of monocytic myeloid-derived suppressor cells in the peripheral blood of patients with B-cell chronic lymphocytic leukemia.Materials and methods. The number of myeloid-derived suppressor cells with CD14+HLA-DRlow/–-phenotype was determined in the peripheral blood of patients with B-cell chronic lymphocytic leukemia and healthy donors by flow cytometry. The measurement was carried out at two points, which differed in the concentration of anti-HLA-DR antibodies − 15 and 4 μl.Results. The median amount of myeloid-derived suppressor cells in the peripheral blood of B-cell chronic lymphocytic leukemia patients with 15 μl of anti-HLA-DR antibody was 1.9 %, and with 4 μl of antibody concentration – 7 %. Healthy donors had that median of 0.15 % with 15 μl of antibody and 0.3 % with 4 μl concentration.Conclusion. The number of CD14+HLA-DRlow/–-cells in the blood of patients with B-cell chronic lymphocytic leukemia is sensitive to the concentration of the HLA-DR antibody.Compliance with patient rights and principles of bioethics. The study protocol No 3 of 13.02.2020 was approved by the biomedical ethics committee of Research Institute of Experimental Diagnostics and Therapy of Tumors N.N. Blokhin National Medical Research Center of Oncology of the Ministry of Health of the Russian Federation. All patients gave written informed consent to participate in the study.

Mesenchymal STEM CELLS Favor Tumor Growth By Generating Granulocyte-like Myeloid Derived Suppressor CELLS in CML Patients

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4018-4018
Author(s):  
Cesarina Giallongo ◽  
Nunziatina L Parrinello ◽  
Daniele Tibullo ◽  
Claudia Bellofiore ◽  
Piera La Cava ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Proliferation ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Suppressor Cells ◽  
T Cell Proliferation ◽  
Myeloid Derived Suppressor Cells ◽  
Healthy Donors

Abstract INTRODUCTION. The complex interplay between cancer cells and immune system allows neoplastic cells to evade immune surveillance and expand. Recently, our and another group have demonstrated that a subpopulation of myeloid cells, defined as "granulocytic myeloid-derived suppressor cells" (G-MDSC), plays an important role for immune escape in chronic myeloid leukemia (CML) patients by reducing T cell activation. The aim of this study was to evaluate the influence of Mesenchymal stem cells (MSC) on generation of MDSCs by comparing CML MSCs (n=10) with healthy donors (HD) MSC (n=8). METHODS. G-MDSC (CD11b+CD33+CD14-HLADR- cells) were analyzed in peripheral blood (PB) of 20 healthy donors (HD) and 30 CML patients at diagnosis by cytofluorimetric analysis. Immuno-suppressive activity was tested through incubation of G-MDSC with autologous CFSE-labeled T cells and stimulation with phytohaemagglutinin (PHA). Controls included a positive T cell proliferation control (T cells plus PHA) and a negative one (T cells only). After three days, T cell proliferation was analyzed by flow cytometry. For G-MDSC generation, human peripheral blood mononucleated cells (PBMC) from HD were cultured alone and with MSC of CML (n=10) or HD (n=8) (1:100 ratio). After one week, G-MDSC were isolated using anti-CD66b magnetic microbeads and the phenotype was confirmed by cytofluorimetric analysis. Expression of ARG1, NOS2, PTGS2, TNFα, TGFβ, IL6, IL10, IL1β was also evaluated using real time PCR. RESULTS. Percentage of cells with a G-MDSC phenotype was greater in PB obtained from CML patients than HD (82.5±9.6% vs 56,2±5.4%, p<0.0001). G-MDSC were able to inhibit T cell proliferation compared to positive control (25±5% vs 48±7.6%, p=0.0057). To investigate if CML MSC may be involved in G-MDSC generation, we incubated HD PBMC with CML or HD MSC for one week. After magnetic isolation, we found that only CML MSC-educated G-MDSC acquired immune-suppressive ability, inhibiting T cell proliferation compared to G-MDSCs control (isolated from PBMC cultured in medium alone) (32±12% vs 63±5.9%, p=0.003). On the contrary, HD MSC-educated G-MDSC did not show any suppressive effect. We also found that CML MSC-educated G-MDSC expressed higher level of the following immune modulatory factors: TNFα (20.8±19.3, p=0.006), IL1β (47.3±25.2, p=0.001), PTGS2 (20.7±10.9, p=0.002) and IL6 (33.8±13.9, p=0.004) compared to HD MSC-educated G-MDSCs (arbitrarily 2-ΔΔCt value: 1). MSC WE also observed ane an up-regulation of PTGS2 (19±4.4, p=0.04), TGFβ (6±3, p=0.01) and IL6 (5±2.8, p=0.04) in CML MSCs at time 0 with a great variability among the patients (calculated value of 2-ΔΔCt in HD MSC was 1). After 48 h of co-culture with PBMC, CML MSC showed statistically significant up-regulation of ARG1 (23.5±11.9, p=0.02), TGFβ (4.8±3, p=0.04), IL10 (5.6±2.8, p=0.03) and IL6 (54.3±23, p=0.02) expression, suggesting that multiple mechanisms are involved in MDSC induction by CML MSC. CONCLUSION. Our work demonstrates that CML MSCs are able to activate MDSCs favoring cancer immune evasion in CML patients. Disclosures Palumbo: Novartis: Honoraria, Other: Advisory Board.

scholarly journals P862 Clinical benefit potentially evident with immunopharmacodynamic responses in prior-checkpoint failed metastatic melanoma patients treated with imprime PGG and pembrolizumab

2020 ◽  
Vol 8 (Suppl 1) ◽  
pp. A10.1-A10
Author(s):  
Anissa Chan ◽  
Nandita Bose ◽  
Nandita Bose ◽  
Nadine Ottoson ◽  
Xiaohong Qiu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Disease Control ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Clinical Benefit ◽  
Secondary Resistance ◽  
Enhanced Production ◽  
Hla Dr ◽  
Melanoma Patients

BackgroundCheckpoint inhibitor (CPI) monotherapy has revolutionized the treatment of melanoma, yet most patients are primary nonresponders or develop secondary resistance. Lack of antigen-specific T cell priming and/or immunosuppressive mechanisms leading to T cell exhaustion are critical cancer-extrinsic factors contributing to CPI resistance mechanisms. Immunotherapeutic agents capable of sparking de novo anti-tumor T cell responses or reinvigorating pre-existing exhausted T cell immunity could help reinstate the activity of CPI.MethodsOur Phase 2, multi-center, open label study, NCT02981303 in collaboration with Merck & Co., Inc., is evaluating Imprime PGG (Imprime), a novel yeast derived, Dectin-1 agonist, β-glucan PAMP in combination with pembrolizumab (KEYTRUDA®, pembro) in heavily CPI pre-treated melanoma patients (20 patients; 65% had >2 prior CPI regimens with 17/20 having previously progressed on pembro). Patients received Imprime (4 mg/kg) + pembro (200 mg) intravenously in a 3-week cycle. Here, we present the immunopharmacodynamic (IPD) responses elicited by Imprime and pembro in the peripheral blood of 19 patients.ResultsIn the intent-to-treat population (ITT; N=20), the disease control rate was 45% (1 CR and 8 SD), 6-month and 12-month OS rates were 65% and 45% respectively, and median OS (mOS) was 8.8 months. In the patients showing disease control, a significant increase in CH50, the classical pathway complement function (~0.7-2.6-fold), HLA-DR expression on classical monocytes (~0.61-1.94-fold) and reduction of frequency of PD-1+Tbet-EOMES+ exhausted CD8 T cells (~0.9-4-fold) was observed. Stimulation of peripheral blood mononuclear cells from a subset of patients by CD3/CD28 beads showed enhanced production of IL-2 and IFN-gamma in the CD8 T cells. Some of these IPD responses were also associated with 6-month landmark OS analyses. Additionally, whole blood gene expression analyses showed >2-fold upregulation of several myeloid and T cell activation genes including IFNg, CD83, IP-10, and IL-2RA. Enhanced OS was observed in patients with >1.3 fold increase in CH50 (8/19; HR 0.385; p=0.1) or >1.5-fold reduction in the frequency of exhausted CD8 T cells (8/19; HR 0.102; p=0.001). The IPD responses observed in the ITT population included formation of circulating immune complexes (peak levels ranging from ~4.5-16.1-fold) and production of complement activation protein SC5b9 (~3.4-25.6-fold), and increase in the frequency of HLA-DR+ myeloid cells (~0.43-3.71-fold).ConclusionsOverall, these data, albeit in a small population, demonstrate that Imprime/pembro combination can drive the innate/adaptive IPD responses that are critical for providing clinical benefit to the patients who have progressed through prior CPI treatments.Ethics ApprovalThe study was approved by central and local ethics committees depending on site requirements. The central IRB for the study is Western Institutional Review Board (WIRB), approval number 20162506; all sites received IRB approval before opening the study at the respective sites.

CD3+CD4low and CD3+CD8low are induced by HLA-G: novel human peripheral blood suppressor T-cell subsets involved in transplant acceptance

Blood ◽  
2007 ◽  
Vol 110 (12) ◽  
pp. 3936-3948 ◽  
Author(s):  
Abderrahim Naji ◽  
Solene Le Rond ◽  
Antoine Durrbach ◽  
Irene Krawice-Radanne ◽  
Caroline Creput ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Plasma Concentrations ◽  
Suppressor Cells ◽  
Peripheral Tolerance ◽  
T Cell Subsets ◽  
Hla Dr ◽  
Antigen Presenting

Abstract HLA-G is a tolerogenic molecule whose detection in sera and within allografted tissues is associated with better graft acceptance. HLA-G mediates T-cell differentiation into suppressor cells, which are thought to promote tolerance. Here, we investigated such T cells phenotypically and functionally and assessed their clinical relevance in the peripheral blood of patients who have undergone transplantation. Our results demonstrate that HLA-G expressed by antigen-presenting cells or present as soluble protein down-regulates the expression of CD4 and CD8 on allostimulated T cells at both transcriptional and posttranslational levels. These CD3+CD4low and CD3+CD8low T-cell subsets are characterized by an increased proportion of cells expressing CD45RA and HLA-DR, and a decreased number of cells expressing CD62L. In addition, these HLA-G–induced CD3+CD4low and CD3+CD8low subpopulations are Foxp3-negative suppressor T cells whose function involves IL-10. Biologic relevance came from analysis of patients who underwent transplantation, with high HLA-G plasma concentrations associated with better graft survival. Peripheral blood from these patients contains increased levels of IL-10 concomitantly to an enhanced representation of CD3+CD4low and CD3+CD8low T cells compared with HLA-G–negative patients who underwent transplantation and healthy individuals. These data define novel immunosuppressive subpopulations of peripheral blood T cells induced by HLA-G with potent implications in peripheral tolerance.

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