scholarly journals CyTOF Analysis Reveals a Distinct Immunosuppressive Microenvironment in IDH Mutant Anaplastic Gliomas

2021 ◽  
Vol 10 ◽  
Author(s):  
Weilun Fu ◽  
Wenjing Wang ◽  
Hao Li ◽  
Yuming Jiao ◽  
Jiancong Weng ◽  
...  
Keyword(s):  
T Cells ◽  
Mononuclear Cells ◽  
Mononuclear Phagocytes ◽  
High Dimensional ◽  
Mass Cytometry ◽  
Immune Microenvironment ◽  
Peripheral Blood Mononuclear ◽  
Anaplastic Gliomas ◽  
Immunosuppressive Microenvironment ◽  
Immune Changes

The immune microenvironment is important for the development, progression, and prognosis of anaplastic glioma (AG). This complex milieu has not been fully elucidated, and a high-dimensional analysis is urgently required. Utilizing mass cytometry (CyTOF), we performed an analysis of immune cells from 5 patients with anaplastic astrocytoma, IDH-mutant (AAmut) and 10 patients with anaplastic oligodendroglioma, IDH-mutant and 1p/19q codeletion (AOD) and their paired peripheral blood mononuclear cells (PBMCs). Based on a panel of 33 biomarkers, we demonstrated the tumor-driven immune changes in the AG immune microenvironment. Our study confirmed that mononuclear phagocytes and T cells are the most abundant immunocytes in the AG immune microenvironment. Glioma-associated microglia/macrophages in both AAmut and AOD samples showed highly immunosuppressive characteristics. Compared to those in the PBMCs, the ratios of immune checkpoint-positive exhausted CD4+ T cells and CD8+ T cells were higher at the AG tumor sites. The AAmut immune milieu exhibits more immunosuppressive characteristics than that in AOD.

scholarly journals IMMU-30. UTILIZING A NOVEL MASS CYTOMETRY-BASED IMMUNOMONITORING PLATFORM FOR THE CHARACTERIZATION OF VACCINE-REACTIVE, EPITOPE-SPECIFIC CD8+ T-CELLS IN HLA-A*0201+ PATIENTS WITH K27M+ DIFFUSE MIDLINE GLIOMAS

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi125-vi125
Author(s):  
Jared Taitt ◽  
Payal Watchmaker ◽  
Takahide Nejo ◽  
Neil Almeida ◽  
Kaori Okada ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Time Course ◽  
Mononuclear Cells ◽  
Expression Profiles ◽  
Peptide Vaccine ◽  
Effector Memory ◽  
Mass Cytometry ◽  
Peripheral Blood Mononuclear

Abstract Diffuse midline glioma (DMG), including diffuse intrinsic pontine glioma (DIPG) constitutes up to 20% of pediatric brain cancer and has a median survival of less than one year. We have identified a novel HLA-A*02:01-restricted neoantigen epitope encompassing the H3.3K27M mutation and implemented a pilot clinical trial through the Pacific Pediatric Neuro-Oncology Consortium (PNOC007). Newly diagnosed DIPG patients who are HLA-A2+ and H3.3K27M+ underwent radiation therapy, and then received the H3.3K27M peptide vaccine and tetanus toxoid (TT) peptide emulsified in Montanide in combination with poly-ICLC every 3 weeks for a total of 24 weeks. Our objective is to characterize vaccine-induced H3.3K27M-specific T-cell subpopulations in peripheral blood mononuclear cells through the evaluation of surface markers correlated with activation, memory, and exhaustion phenotypes utilizing a novel H3.3K27M-specific dextramer-based mass cytometry method. Through this approach, the temporal expansion of vaccine-reactive CD8+ T-cells was observed in all of patients (n = 4) who completed a minimum of 18 weeks on the study. These T-cells were subsequently stratified into discrete clusters on a tSNE plot using canonical CD8+ T-cell markers. Resultant clusters were further classified by their expression profiles, revealing distinct effector memory and exhausted subpopulations. Chronological monitoring of these groups indicates the time course-dependent development and persistence of vaccine-reactive exhausted and effector memory CD8+ T-cells in 75% of patients analyzed. Furthermore, a comparative analysis of myeloid subpopulations revealed an inverse correlation between the expansion of monocytic myeloid-derived suppressor cells (M-MDSCs) and length of enrollment in the trial. Future plans include the analysis of regulatory T-cells (Tregs) and MDSCs of all enrolled patients to solidify the relationship between the length of stay on the study and prevalence of immunosuppressive populations. This methodology offers insight into the progression of vaccine-induced patient immune responses and exhibits promise as a platform that may be extrapolated to other immunotherapies.

scholarly journals IMMU-23. A NOVEL MASS CYTOMETRY-BASED MULTI-PARAMETER CHARACTERIZATION OF NEOANTIGEN-REACTIVE CD8+ T-CELLS IN PATIENTS PARTICIPATING IN PNOC007 H3.3K27M PEPTIDE VACCINE CLINICAL TRIAL

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii364-iii365
Author(s):  
Jared Taitt ◽  
Takahide Nejo ◽  
Payal Watchmaker ◽  
Neil Almeida ◽  
Kaori Okada ◽  
...  
Keyword(s):  
Clinical Trial ◽  
T Cells ◽  
Clinical Outcomes ◽  
Cd8 T Cells ◽  
Mononuclear Cells ◽  
Expression Profiles ◽  
Peptide Vaccine ◽  
Effector Memory ◽  
Mass Cytometry ◽  
Peripheral Blood Mononuclear

Abstract BACKGROUND We have identified an HLA-A*02:01-restricted neoantigen epitope encompassing the H3.3K27M mutation and implemented a multi-center clinical trial of the peptide vaccine through the Pacific Pediatric Neuro-Oncology Consortium (PNOC007) for patients with diffuse midline glioma (DMG), including diffuse intrinsic pontine glioma (DIPG). We sought to characterize vaccine-reactive CD8+T-cells subpopulations using their precise activation and developmental status to find their associations with clinical outcomes. METHODS Mass cytometry (CyTOF) analysis was performed on patient-derived peripheral blood mononuclear cells collected at baseline as well as pre-specified time points throughout the study. Each cell subtype was characterized via tSNE-clustering based on their expression profiles and quantified as a fraction of total CD45+cells. H3.3K27M-reactive CD8+T-cells were evaluated using an H3.3K27M-HLA-A2 dextramer along with a panel of T-cell and myeloid markers. RESULTS Among all 29 patients enrolled, we analyzed samples from all 19 DIPG and 9 of 10 non-brainstem DMG cases, of which 18 had longitudinal samples available (range: 2–5). Utilizing a novel CyTOF-based immuno-monitoring platform, the expansion of H3.3K27M-reactive CD8+T-cells, defined as a 25% increase at any time-point relative to baseline, was observed in 7 of these 18 patients. Survival analyses indicated that the expansion of H3.3K27M-reactive CD8+T-cells, particularly the effector-memory phenotype, positively correlated with longer overall survival (OS) (median: 16.1 vs 9.7 months, p=0.03), whereas an abundance of early and monocytic myeloid-derived suppressor cells at baseline correlated with shorter OS among DIPG patients (9.5 vs 14.3 months, p=0.002). CONCLUSION Our novel immuno-monitoring approach offers insight into how vaccine-induced immune responses impact clinical outcomes.

scholarly journals Multi-parameter immune profiling of peripheral blood mononuclear cells by multiplexed single-cell mass cytometry in patients with early multiple sclerosis

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Chotima Böttcher ◽  
Camila Fernández-Zapata ◽  
Stephan Schlickeiser ◽  
Desiree Kunkel ◽  
Axel R. Schulz ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Cell Mass ◽  
Healthy Controls ◽  
Mass Cytometry ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

AbstractMultiple sclerosis (MS) is an inflammatory demyelinating and neurodegenerative disease of the central nervous system (CNS). Studies in rodent models demonstrated an association of CNS-infiltrating monocyte-derived macrophages with disease severity. However, little is known about humans. Here, we performed an exploratory analysis of peripheral blood mononuclear cells (PBMCs) isolated from healthy controls and drug-naïve patients with early MS using multiplexed single-cell mass cytometry and algorithm-based data analysis. Two antibody panels comprising a total of 64 antibodies were designed to comprehensively analyse diverse immune cell populations, with particular emphasis on monocytes. PBMC composition and marker expression were overall similar between the groups. However, an increased abundance of CCR7+ and IL-6+ T cells was detected in early MS-PBMCs, whereas NFAT1hiT-bethiCD4+ T cells were decreased. Similarly, we detected changes in the subset composition of the CCR7+ and MIPβhi HLA-DR+ lymphocyte compartment. Only mild alterations were detected in monocytes/myeloid cells of patients with early MS, namely a decreased abundance of CD141hiIRF8hiCXCR3+CD68− dendritic cells. Unlike in Crohn’s disease, no significant differences were found in the monocyte fraction of patients with early MS compared to healthy controls. This study provides a valuable resource for future studies designed to characterise and target diverse PBMC subsets in MS.

Multidimensional flow characterization of circulating immune cells in cisplatin-resistant metastatic urothelial cancer (mUC) patients (pts) treated with pembrolizumab (P) with or without acalabrutinib (acala).

2019 ◽  
Vol 37 (7_suppl) ◽  
pp. 357-357
Author(s):  
Tian Zhang ◽  
Janet S. Staats ◽  
Cliburn Chan ◽  
Ajjai Shivaram Alva ◽  
Peter H. O'Donnell ◽  
...  
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Mononuclear Cells ◽  
Checkpoint Inhibitors ◽  
Randomized Study ◽  
T Cell Subsets ◽  
Peripheral Blood Mononuclear ◽  
Hla Dr ◽  
Metastatic Urothelial Cancer ◽  
Immune Changes

357 Background: Immunotherapy checkpoint inhibitors (ICIs) are now standard treatments (txs) in mUC, with response rates ranging 15-25%. A phase 2 randomized study (NCT02351739) treated 75 pts with mUC with either P or PA. We profiled peripheral blood mononuclear cells (PBMCs) to understand changes on tx. Methods: Comprehensive immune profiling of PBMCs was performed at baseline, weeks (wk) 4, 7, and 10. The Wilcoxon rank sum (WRS) test was used to compare tx arms, as well as pts with complete and partial response (CR/PR) vs disease progression (PD). Using the Benjamini-Hochberg (BH) method, results were deemed significant if they had false discovery rate (FDR) of 0.1 or less. Results: Clinical responses did not differ between tx arms (Zhang T et al, ESMO, 2017). The majority of patients showed largest immune changes from baseline at wk7. Comparing P vs PA pts, the PA group had statistically significant increases in PD-L1+ monocytes; CD8 T cells expressing: CD39, CTLA-4, ICOS, PD-1, TIM3, HLA-DR, Ki67, CD39/HLA-DR, CD39/Ki67, and PD-1/TIM-3; and exhausted (CD28-) CD8 T cells expressing ICOS and PD-L1 (Table). Comparing CR/PR vs PD pts, Treg subsets expressing CCR4/HLA-DR and CD39/HLA-DR were significant by WRS but not FDR criteria. Conclusions: From this extensive, exploratory, non-functional flow analysis of PBMCs, PA may potentiate immune changes of PD-L1+ monocytes and CD8+ T cell subsets. Increasing activated peripheral Tregs in CR/PR pts in either arm may indicate marginalization of Tregs from the tumor microenvironment. [Table: see text]

scholarly journals Alterations in the phenotype of hairy cells during culture in the presence of PHA: requirement for T cells

Blood ◽  
1982 ◽  
Vol 59 (5) ◽  
pp. 895-899 ◽  
Author(s):  
CP Worman ◽  
PC Beverley ◽  
JC Cawley
Keyword(s):  
T Cells ◽  
Hairy Cell Leukemia ◽  
Mononuclear Cells ◽  
Cultured Cells ◽  
Phenotypic Change ◽  
Cell Contact ◽  
Hairy Cell ◽  
Cell Leukemia ◽  
Peripheral Blood Mononuclear ◽  
Hairy Cells

Abstract Culture studies of peripheral blood mononuclear cells from 7 entirely typical cases of hairy cell leukemia showed that after culture in the presence of PHA for 2--5 days, the predominant cell type changed from E- SIg+ CIg+ gamma FcR+ muFcR+ hairy cells to an E+ SIg- CIg- gamma FcR- muFcR- population of transformed cells derived from hairy cells. Depletion and readdition experiments demonstrated that cell-to-cell contact with T cells was necessary for the phenotypic change, while several observations indicated that the E+ population was not derived from T cells present before culture. The E positivity of the cultured cells was shown to be due to the possession of E receptor not acquired from the culture fluid, but the cells differed from true T cells in lacking both mature and immature T-cell antigens. The relevance of these in vitro observations to the continuing controversy concerning the nature of the hairy cell and to the in vivo fluctuations in immunologic phenotype not infrequently observed in hairy cell leukemia is briefly discussed.

scholarly journals Deconvolution of monocyte responses in inflammatory bowel disease reveals an IL-1 cytokine network that regulates IL-23 in genetic and acquired IL-10 resistance

Gut ◽  
2020 ◽  
pp. gutjnl-2020-321731
Author(s):  
Dominik Aschenbrenner ◽  
Maria Quaranta ◽  
Soumya Banerjee ◽  
Nicholas Ilott ◽  
Joanneke Jansen ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Personalised Medicine ◽  
Mononuclear Phagocytes ◽  
Cytokine Network ◽  
Peripheral Blood Mononuclear ◽  
Transcriptional Signature ◽  
Inflammatory Monocytes ◽  
Transcriptomic Signature ◽  
Inflammatory Bowel

ObjectiveDysregulated immune responses are the cause of IBDs. Studies in mice and humans suggest a central role of interleukin (IL)-23-producing mononuclear phagocytes in disease pathogenesis. Mechanistic insights into the regulation of IL-23 are prerequisite for selective IL-23 targeting therapies as part of personalised medicine.DesignWe performed transcriptomic analysis to investigate IL-23 expression in human mononuclear phagocytes and peripheral blood mononuclear cells. We investigated the regulation of IL-23 expression and used single-cell RNA sequencing to derive a transcriptomic signature of hyperinflammatory monocytes. Using gene network correlation analysis, we deconvolved this signature into components associated with homeostasis and inflammation in patient biopsy samples.ResultsWe characterised monocyte subsets of healthy individuals and patients with IBD that express IL-23. We identified autosensing and paracrine sensing of IL-1α/IL-1β and IL-10 as key cytokines that control IL-23-producing monocytes. Whereas Mendelian genetic defects in IL-10 receptor signalling induced IL-23 secretion after lipopolysaccharide stimulation, whole bacteria exposure induced IL-23 production in controls via acquired IL-10 signalling resistance. We found a transcriptional signature of IL-23-producing inflammatory monocytes that predicted both disease and resistance to antitumour necrosis factor (TNF) therapy and differentiated that from an IL-23-associated lymphocyte differentiation signature that was present in homeostasis and in disease.ConclusionOur work identifies IL-10 and IL-1 as critical regulators of monocyte IL-23 production. We differentiate homeostatic IL-23 production from hyperinflammation-associated IL-23 production in patients with severe ulcerating active Crohn’s disease and anti-TNF treatment non-responsiveness. Altogether, we identify subgroups of patients with IBD that might benefit from IL-23p19 and/or IL-1α/IL-1β-targeting therapies upstream of IL-23.

Class I–unrestricted noncytotoxic anti–HTLV-I activity of CD8+ T cells

Blood ◽  
2000 ◽  
Vol 96 (5) ◽  
pp. 1994-1995 ◽  
Author(s):  
Masako Moriuchi ◽  
Hiroyuki Moriuchi
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Mononuclear Cells ◽  
Membrane Filter ◽  
Cd8 T Cell ◽  
Class I ◽  
Type I ◽  
Peripheral Blood Mononuclear ◽  
Permeable Membrane

Abstract Although it is widely believed that viral clearance is mediated principally by the destruction of infected cells by cytotoxic T cells, noncytolytic antiviral activity of CD8+ T cells may play a role in preventing the progression to disease in infections with immunodeficiency viruses and hepatitis B virus. We demonstrate here that (1) replication of human T-lymphotropic virus type I (HTLV-I) is more readily detected from CD8+ T-cell–depleted (CD8−) peripheral blood mononuclear cells (PBMCs) of healthy HTLV-I carriers than from unfractionated PBMCs, (2) cocultures of CD8− PBMCs with autologous or allogeneic CD8+ T cells suppressed HTLV-I replication, and (3) CD8+ T-cell anti-HTLV-I activity is not abrogated intrans-well cultures in which CD8+ cells are separated from CD8− PBMCs by a permeable membrane filter. These results suggest that class I-unrestricted noncytolytic anti–HTLV-I activity is mediated, at least in part by a soluble factor(s), and may play a role in the pathogenesis of HTLV-I infection.

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