PD-1+ T Cell Subsets in Follicular Lymphoma Tumor Microenvironment and Their Implications for Prognosis and Therapy

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2648-2648
Author(s):  
Fuliang Chu ◽  
Wencai Ma ◽  
Tomohide Yamazaki ◽  
Myriam Foglietta ◽  
Durga Nattama ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Follicular Lymphoma ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
T Cell Subsets ◽  
Prognostic Impact ◽  
T Cell Exhaustion ◽  
Cell Exhaustion

Abstract Abstract 2648 Background: Programmed death (PD)-1, a coinhibitory receptor expressed by effector T cells (Teffs) is highly expressed on intratumoral T cells (mean 61%, range 34–86% for CD4+ T cells and mean 44%, range 31–69% for CD8+ T cells) in follicular lymphoma (FL), a finding associated with impaired ability to recognize autologous tumor (Nattamai et al, ASH 2007). Hence, PD-1 expression would be expected to confer an unfavorable prognosis in FL. However, correlation of PD-1 with clinical outcome in FL has been inconsistent with two studies showing favorable (Carreras et al, J Clin Oncol 2009; Wahlin et al, Clin Cancer Res 2010) and one study showing unfavorable (Richendollar et al, Hum Pathol 2011) outcome. While differences in method of analysis and type of treatment may explain the disparate results, a more complex model may be necessary to understand the prognostic impact of PD-1 in FL as PD-1 is expressed not only on antitumor Teffs but also on protumor follicular helper T cells (Tfh) and regulatory T cells (Tregs). Methods: To determine the nature of PD-1+ T cells in FL we performed comprehensive genomic and immunologic studies. By flow cytometry, we observed that the intratumoral CD4+ T cells in FL may be categorized into 3 subsets based on PD-1 expression - PD-1 high (PD-1hi), intermediate (PD-1int), and low (PD-1lo). The intratumoral CD8+ T cells consisted of PD-1int and PD-1lo subsets. The 3 CD4+ T cell subsets were FACSorted from FL tumors (n=3) and whole genome gene expression profiling (GEP) was performed. T cell subsets sorted similarly from tonsils served as controls for reactive follicular hyperplasia (FH) (n=3). Differentially expressed genes in GEP studies were confirmed at the mRNA level by real-time PCR (n=5) and at the protein level by flow cytometry when antibodies were available (n=5–10). Results: Our results suggested that CD4+PD-1hi T cells are Tfh cells (CXCR5hiBcl6hi ICOShiCD40LhiSAPhiPRDM1loIL-4hiIL-21hi); the CD4+PD-1int T cells consisted of a mixture of activated Teffs (CD45RO+CD45RA−) including Th1 (Tbet+IFNg+), Th2 (IL-10+), and Th17 cells (RORc+IL-17+), and Tregs (Foxp3+CD25hiCD127lo); and the CD4+PD-1lo T cells consisted of a mixture of activated Teffs (CD45RO+CD45RA− but IFNg−IL-4−IL-10−IL-17−), Tregs, and naïve T cells (CD45RO−CD45RA+CCR7+). Although these subsets were present in both FL and FH, there were important differences. IL-4 expression was significantly higher in Tfh in FL vs. FH and may play a role in the pathogenesis of FL. IL-17 expression was low and expression of coinhibitory molecules BTLA and CD200 was high in CD4+PD-1int T cells in FL vs. FH. BTLA and CD200 were also increased in CD8+PD-1int T cells in FL vs. FH. However, other coinhibitory molecules (LAG-3, Tim-3, CD160, CTLA-4, CD244, KLRG1) were not significantly different between FL and FH. CD4+PD-1int T cells also had higher expression of BATF, a transcription factor associated with T cell exhaustion in FL vs. FH. Together, these results suggest that the CD4+PD-1int T cells in FL may be in a state of T cell exhaustion whereas the CD4+PD-1int T cells in FH may represent recently activated Teffs. Consistent with this, blocking PD-1 with anti-PD-1 blocking antibody significantly enhanced proliferation and the production of Th1 (IFNg, TNFa) but not Th2 (IL-4, IL-5, IL-10, IL-13) cytokines by intratumoral CD4+ and CD8+ T cells in response to stimulation with autologous FL tumor cells (n=3). As expected, Tregs were increased in number in FL vs. FH and were present in the PD-1int and PD-1lo T cell subsets. We found 74% (range 40–97%) of FL Tregs expressed PD-1. Among the CD4+PD-1lo and CD8+PD-1lo T cells, there were more activated Teffs and fewer naïve T cells in FL vs. FH. Conclusions: Our results suggest that the PD-1+ T cells in FL are comprised of a mixture of antitumor Teffs and protumor Tfh and Tregs. The prognostic impact of PD-1+ T cells in FL may dependent on the relative frequency of these subsets as ligation of PD-1 may produce favorable (inhibition of protumor Tfh and Tregs) or unfavorable (inhibition of antitumor Teffs) outcomes by inhibiting or promoting tumor growth, respectively. Conversely, our results imply that agents that block PD-1/PD-ligand pathway may have the opposite effect on these T cell subsets and enumeration of the intratumoral PD-1+ T cell subsets may serve as biomarker to predict response to these agents in FL and possibly other B-cell malignancies. Disclosures: Dong: GSK: Consultancy; Genentech: Honoraria; Tempero: Consultancy; Ono: Consultancy; AnaptysBio: Consultancy. Neelapu:Cure Tech Ltd: Research Funding.

scholarly journals 644 Tumor-mediated suppressive signaling and hypoxia supports altered chromatin landscapes that limit the transcriptional and functional potential of terminal T cell exhaustion

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A673-A673
Author(s):  
Rhodes Ford ◽  
Natalie Rittenhouse ◽  
Nicole Scharping ◽  
Paolo Vignali ◽  
Greg Delgoffe ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Histone Modifications ◽  
Cd8 T Cells ◽  
Tumor Hypoxia ◽  
T Cell Subsets ◽  
T Cell Exhaustion ◽  
Cell Exhaustion

BackgroundCD8+ T cells are a fundamental component of the anti-tumor response; however, tumor-infiltrating CD8+ T cells (TIL) are rendered dysfunctional by the tumor microenvironment. CD8+ TIL display an exhausted phenotype with decreased cytokine expression and increased expression of co-inhibitory receptors (IRs), such as PD-1 and Tim-3. The acquisition of IRs mark the progression of dysfunctional TIL from progenitors (PD-1Low) to terminally exhausted (PD-1+Tim-3+). How the chromatin landscape changes during this progression has not been described.MethodsUsing a low-input ChIP-based assay called Cleavage Under Targets and Release Using Nuclease (CUT&RUN), we have profiled the histone modifications at the chromatin of tumor-infiltrating CD8+ T cell subsets to better understand the relationship between the epigenome and the transcriptome as TIL progress towards terminal exhaustion.ResultsWe have identified two epigenetic characteristics unique to terminally exhausted cells. First, we have identified a unique set of genes, characterized by active histone modifications that do not have correlated gene expression. These regions are enriched for AP-1 transcription factor motifs, yet most AP-1 family factors are actively downregulated in terminally exhausted cells, suggesting signals that promote downregulation of AP-1 expression negatively impacts gene expression. We have shown that inducing expression of AP-1 factors with a 41BB agonist correlates with increased expression of these anticorrelated genes. We have also found a substantial increase in the number of genes that exhibit bivalent chromatin marks, defined by the presence of both active (H3K4me3) and repressive (H3K27me3) chromatin modifications that inhibit gene expression. These bivalent genes in terminally exhausted T cells are not associated with plasticity and represent aberrant hypermethylation in response to tumor hypoxia, which is necessary and sufficient to promote downregulation of bivalent genes.ConclusionsOur study defines for the first time the roles of costimulation and the tumor microenvironment in driving epigenetic features of terminally exhausted tumor-infiltrating T cells. These results suggest that terminally exhausted T cells have genes that are primed for expression, given the right signals and are the basis for future work that will elucidate that factors that drive progression towards terminal T cell exhaustion at the epigenetic level and identify novel therapeutic targets to restore effector function of tumor T cells and mediate tumor clearance.

scholarly journals The expression of class II major histocompatibility molecules on breast tumors delays T cell exhaustion, expands the T cell repertoire and slows tumor growth

2018 ◽  
Author(s):  
Tyler R. McCaw ◽  
Mei Li ◽  
Dmytro Starenki ◽  
Sara J. Cooper ◽  
Selene Meza-Perez ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Tumor Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
T Cell Exhaustion ◽  
Major Histocompatibility ◽  
Cell Exhaustion ◽  
Mhcii Expression

AbstractThe expression of major histocompatibility complex II (MHCII) on tumor cells correlates with survival and responsiveness to immunotherapy. However, the mechanisms underlying these observations are poorly defined. Using a murine breast tumor line, we tested how MHCII expression affected anti-tumor immunity. We found that MHCII-expressing tumors grew more slowly than controls and recruited more functional CD4+ and CD8+ T cells. Additionally, MHCII-expressing tumors contained more TCR clonotypes expanded to a larger degree than control tumors. Functional CD8+ T cells in tumors depended on CD4+ T cells. However, both CD4+ and CD8+ T cells eventually became exhausted, even in MHCII-expressing tumors. PD1 blockade had no impact on tumor growth, potentially because tumor cells poorly expressed PD-L1. These results suggest tumor cell expression of MHCII facilitates the local activation of CD4+ T cells and indirectly helps the activation and expansion of CD8+ T cells, but by itself, cannot prevent T cell exhaustion.PrécisThe expression of MHCII on tumor cells augments CD4 and CD8 T cell responses, expands the TCR repertoire and delays exhaustion. Hence, strategies to induce MHCII expression may be a powerful adjuvant to immunotherapeutic regimens of solid tumors.

scholarly journals CLL Is Associated with Development of Subclinical Cytomegalovirus Viraemia and Accumulation of Large Populations of “exhausted” CMV-Specific CD4+ T Cells

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3290-3290
Author(s):  
Helen M Parry ◽  
Natasha Cutmore ◽  
Nikhil Mirajkar ◽  
Annette Pachnio ◽  
Tina McSkeane ◽  
...  
Keyword(s):  
T Cells ◽  
Viral Load ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Class Ii ◽  
Cd4 T Cell ◽  
Effector Memory ◽  
T Cell Exhaustion ◽  
Cell Exhaustion

Abstract Chronic Lymphocytic Leukaemia (CLL) is associated with T cell dysfunction and increased expression of markers of T cell exhaustion. Cytomegalovirus is a common herpesvirus infection and is associated with development of accelerated immune senescence in older adults. Within patients with CLL, CMV leads to marked expansion of virus-specific CD4 and CD8 T cells and development of oligoclonal T cell populations. However these features are not known to be associated with clinical symptoms and CMV viral load remains essentially undetectable by conventional qPCR. In this study we used digital PCR to determine CMV viral load in patients with CLL and correlated this with the magnitude and phenotype of the CMV-specific T cell immune response. In particular, we utilised HLA class II tetramers for the first time, in order to assess the contribution of CMV-specific T cell populations to the profile of T cell exhaustion seen in this disease. 68 CMV-seropositive CLL patients and 19 age-matched healthy donors (HD) were recruited for study. All patients were either untreated or had not received chemotherapy for at least 6 months. CMV viral load was determined within purified monocyte populations using a digital droplet PCR method. Viral load per monocyte was increased in CLL patients compared to HD (p=0.04), with the highest viral loads detected in stage C patients. In order to investigate the CMV-specific immune response, nine HLA class I tetramers were generated, containing viral epitopes from pp65, pp50 and IE-1, and two class II tetramers were also available, with epitopes from glycoprotein B and pp65. CMV-specific CD4 T cell responses were increased in CLL patients (n=14) compared to HD (n=11) (4.1 % vs. 0.9 %; p=0.049). Remarkably, in one patient more than half (50.9%) of all CD4 T cells were directed against a single CMV epitope. CD4 CMV-specific T cells were nearly always effector memory in phenotype (78%), somewhat in contrast to CD8 populations where the memory phenotype is split between effector memory (CCR7-, CD45RA-) and terminally differentiated memory cells (CCR7-, CD45RA+). PD1 is an important marker of T cell exhaustion and expression was increased on both CD8 T cells (16.9% Vs 9.6%; p=0.003); and CD4 T cells (16.2 % Vs 8.7 %; p=0.0007) in CLL patients compared to HD. Interestingly, PD1 expression was much higher on CMV-specific CD4 T cells compared to the total CD4 T cell repertoire (50.6% Vs 21%; p=0.01), whereas the opposite profile was observed in relation to CD8 populations. CMV-specific CD4 T cells demonstrated a Th1 cytotoxic phenotype with production of TNF-alpha, IFN-y and Granzyme B. Production of IFN-y and TNF-alpha was reduced in PD1+ populations, consistent with an exhausted phenotype. No CD25+FoxP3 + regulatory T cells were observed within the CMV-specific CD4 T cell population which is of interest given the well documented expansion of this population in patients with CLL. In order to investigate the replicative history of CMV-specific T cells we investigated the telomere length of antigen-specific cells using single cell telomere length analysis. CMV-specific cells had markedly shortened telomere lengths compared to background CD4 and CD8 T cells, with the mean difference of 0.911 kb (maximum 1.836 kb) indicating that they have undergone extensive proliferation in vivo. This work is the first to use digital PCR to measure the subclinical CMV load within peripheral blood and also to examine CMV-specific CD4 T cells using HLA class II tetramers. Our results indicate that immune control of CMV viral load is impaired during the clinical progression of CLL and that a proportion of virus-specific CD4 T cells show signs of exhaustion. These data may reflect ‘cross presentation’ of viral protein by B-CLL tumour cells, which are known to have poor capacity for antigen presentation. The potential clinical importance of these observations is now being addressed. Disclosures No relevant conflicts of interest to declare.

scholarly journals Partial restoration of immune response in Hepatitis C patients after viral clearance by direct-acting antiviral therapy

PLoS ONE ◽  
2021 ◽  
Vol 16 (7) ◽  
pp. e0254243
Author(s):  
Meritxell Llorens-Revull ◽  
Maria Isabel Costafreda ◽  
Angie Rico ◽  
Mercedes Guerrero-Murillo ◽  
Maria Eugenia Soria ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
Hepatitis C ◽  
T Cell ◽  
Cd8 T Cells ◽  
Viral Clearance ◽  
Cd4 T Cell ◽  
Care Hospital ◽  
T Cell Exhaustion ◽  
Cell Exhaustion

Background & aims HCV CD4+ and CD8+ specific T cells responses are functionally impaired during chronic hepatitis C infection. DAAs therapies eradicate HCV infection in more than 95% of treated patients. However, the impact of HCV elimination on immune responses remain controversial. Here, we aimed to investigate whether HCV cure by DAAs could reverse the impaired immune response to HCV. Methods We analyzed 27 chronic HCV infected patients undergoing DAA treatment in tertiary care hospital, and we determined the phenotypical and functional changes in both HCV CD8+ and CD4+ specific T-cells before and after viral clearance. PD-1, TIM-3 and LAG-3 cell-surface expression was assessed by flow cytometry to determine CD4+ T cell exhaustion. Functional responses to HCV were analyzed by IFN-Ɣ ELISPOT, intracellular cytokine staining (IL-2 and IFN-Ɣ) and CFSE-based proliferation assays. Results We observed a significant decrease in the expression of PD-1 in CD4+ T-cells after 12 weeks of viral clearance in non-cirrhotic patients (p = 0.033) and in treatment-naive patients (p = 0.010), indicating a partial CD4 phenotype restoration. IFN-Ɣ and IL-2 cytokines production by HCV-specific CD4+ and CD8+ T cells remained impaired upon HCV eradication. Finally, a significant increase of the proliferation capacity of both HCV CD4+ and CD8+ specific T-cells was observed after HCV elimination by DAAs therapies. Conclusions Our results show that in chronically infected patients HCV elimination by DAA treatment lead to partial reversion of CD4+ T cell exhaustion. Moreover, proliferative capacity of HCV-specific CD4+ and CD8+ T cells is recovered after DAA’s therapies.

scholarly journals Single-Cell Transcriptome Analysis Reveals RGS1 as a New Marker and Promoting Factor for T-Cell Exhaustion in Multiple Cancers

2021 ◽  
Vol 12 ◽  
Author(s):  
Yunmeng Bai ◽  
Meiling Hu ◽  
Zixi Chen ◽  
Jinfen Wei ◽  
Hongli Du
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Poor Prognosis ◽  
Effector T Cells ◽  
T Cell Exhaustion ◽  
Cell Exhaustion ◽  
Cell Transcriptome ◽  
Cancer Tissues ◽  
Single Cell Transcriptome

T-cell exhaustion is one of the main reasons of tumor immune escape. Using single-cell transcriptome data of CD8+ T cells in multiple cancers, we identified different cell types, in which Pre_exhaust and exhausted T cells participated in negative regulation of immune system process. By analyzing the coexpression network patterns and differentially expressed genes of Pre_exhaust, exhausted, and effector T cells, we identified 35 genes related to T-cell exhaustion, whose high GSVA scores were associated with significantly poor prognosis in various cancers. In the differentially expressed genes, RGS1 showed the greatest fold change in Pre_exhaust and exhausted cells of three cancers compared with effector T cells, and high expression of RGS1 was also associated with poor prognosis in various cancers. Additionally, RGS1 protein was upregulated significantly in tumor tissues in the immunohistochemistry verification. Furthermore, RGS1 displayed positive correlation with the 35 genes, especially highly correlated with PDCD1, CTLA4, HAVCR2, and TNFRSF9 in CD8+ T cells and cancer tissues, indicating the important roles of RGS1 in CD8+ T-cell exhaustion. Considering the GTP-hydrolysis activity of RGS1 and significantly high mRNA and protein expression in cancer tissues, we speculated that RGS1 potentially mediate the T-cell retention to lead to the persistent antigen stimulation, resulting in T-cell exhaustion. In conclusion, our findings suggest that RGS1 is a new marker and promoting factor for CD8+ T-cell exhaustion and provide theoretical basis for research and immunotherapy of exhausted cells.

scholarly journals Unraveling the Multifaceted Nature of CD8 T Cell Exhaustion Provides the Molecular Basis for Therapeutic T Cell Reconstitution in Chronic Hepatitis B and C

Cells ◽  
2021 ◽  
Vol 10 (10) ◽  
pp. 2563
Author(s):  
Valeria Barili ◽  
Andrea Vecchi ◽  
Marzia Rossi ◽  
Ilaria Montali ◽  
Camilla Tiezzi ◽  
...  
Keyword(s):  
T Cells ◽  
Chronic Hepatitis ◽  
T Cell ◽  
Hepatitis B ◽  
Chronic Hepatitis B ◽  
Cd8 T Cells ◽  
Stress Responses ◽  
T Cell Exhaustion ◽  
Virus Infections ◽  
Cell Exhaustion

In chronic hepatitis B and C virus infections persistently elevated antigen levels drive CD8+ T cells toward a peculiar differentiation state known as T cell exhaustion, which poses crucial constraints to antiviral immunity. Available evidence indicates that T cell exhaustion is associated with a series of metabolic and signaling deregulations and with a very peculiar epigenetic status which all together lead to reduced effector functions. A clear mechanistic network explaining how intracellular metabolic derangements, transcriptional and signaling alterations so far described are interconnected in a comprehensive and unified view of the T cell exhaustion differentiation profile is still lacking. Addressing this issue is of key importance for the development of innovative strategies to boost host immunity in order to achieve viral clearance. This review will discuss the current knowledge in HBV and HCV infections, addressing how innate immunity, metabolic derangements, extensive stress responses and altered epigenetic programs may be targeted to restore functionality and responsiveness of virus-specific CD8 T cells in the context of chronic virus infections.

PD-1 up-regulation is correlated with HIV-specific memory CD8+ T-cell exhaustion in typical progressors but not in long-term nonprogressors

Blood ◽  
2007 ◽  
Vol 109 (11) ◽  
pp. 4671-4678 ◽  
Author(s):  
Ji-Yuan Zhang ◽  
Zheng Zhang ◽  
Xicheng Wang ◽  
Jun-Liang Fu ◽  
Jinxia Yao ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Effector Memory ◽  
T Cell Exhaustion ◽  
Cell Exhaustion ◽  
Specific Memory ◽  
Memory Cd8 T Cell

Abstract The immunoreceptor PD-1 is significantly up-regulated on exhausted CD8+ T cells during chronic viral infections such as HIV-1. However, it remains unknown whether PD-1 expression on CD8+ T cells differs between typical progressors (TPs) and long-term nonprogressors (LTNPs). In this report, we examined PD-1 expression on HIV-specific CD8+ T cells from 63 adults with chronic HIV infection. We found that LTNPs exhibited functional HIV-specific memory CD8+ T cells with markedly lower PD-1 expression. TPs, in contrast, showed significantly up-regulated PD-1 expression that was closely correlated with a reduction in CD4 T-cell number and an elevation in plasma viral load. Importantly, PD-1 up-regulation was also associated with reduced perforin and IFN-γ production, as well as decreased HIV-specific effector memory CD8+ T-cell proliferation in TPs but not LTNPs. Blocking PD-1/PD-L1 interactions efficiently restored HIV-specific CD8+ T-cell effector function and proliferation. Taken together, these findings confirm the hypothesis that high PD-1 up-regulation mediates HIV-specific CD8+ T-cell exhaustion. Blocking the PD-1/PD-L1 pathway may represent a new therapeutic option for this disease and provide more insight into immune pathogenesis in LTNPs.

A Detailed Phenotypic Analysis Using Six Color Flow Cytometry of Lymphocyte Subsets Mobilized with AMD3100 Compared to G-CSF.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 408-408 ◽  
Author(s):  
Yoshiyuki Takahashi ◽  
S. Chakrabarti ◽  
R. Sriniivasan ◽  
A. Lundqvist ◽  
E.J. Read ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Lymphocyte Subsets ◽  
Phenotypic Analysis ◽  
Color Flow ◽  
Cd34 Cells

Abstract AMD3100 (AMD) is a bicyclam compound that rapidly mobilizes hematopoietic progenitor cells into circulation by inhibiting stromal cell derived factor-1 binding to its cognate receptor CXCR4 present on CD34+ cells. Preliminary data in healthy donors and cancer patients show large numbers of CD34+ cells are mobilized following a single injection of AMD3100. To determine whether AMD3100 mobilized cells would be suitable for allografting, we performed a detailed phenotypic analysis using 6 color flow cytometry (CYAN Cytometer MLE) of lymphocyte subsets mobilized following the administration of AMD3100, given as a single 240mcg/kg injection either alone (n=4) or in combination with G-CSF (n=2: G-CSF 10 mcg/kg/day x 5: AMD3100 given on day 4). Baseline peripheral blood (PB) was obtained immediately prior to mobilization; in recipients who received both agents, blood was analyzed 4 days following G-CSF administration as well as 12 hours following administration of AMD3100 and a 5th dose of G-CSF. AMD3100 alone significantly increased from baseline the PB WBC count (2.8 fold), Absolute lymphocyte count (ALC: 2.5 fold), absolute monocyte count (AMC: 3.4 fold), and absolute neutrophil count (ANC: 2.8 fold). Subset analysis showed AMD3100 preferentially increased from baseline PB CD34+ progenitor counts (5.8 fold), followed by CD19+ B-cells (3.7 fold), CD14+ monocytes (3.4 fold), CD8+ T-cells (2.5 fold), CD4+ T-cells (1.8 fold), with a smaller increase in CD3−/CD16+ or CD56+ NK cell counts (1.6 fold). There was no change from baseline in the % of CD4+ or CD8+ T-cell expressing CD45RA, CD45RO, or CD56, CD57, CD27, CD71 or HLA-DR. In contrast, there was a decline compared to baseline in the mean percentage of CD3+/CD4+ T-cells expressing CD25 (5.5% vs 14.8%), CD62L (12.1% vs 41.1%), CCR7 (2.1% vs 10.5%) and CXCR4 (0.5% vs 40.9%) after AMD3100 administration; similar declines in expression of the same 4 surface markers were also observed in CD3+/CD8+ T-cells. A synergistic effect on the mobilization of CD34+ progenitors, CD19+ B cells, CD3+ T-cells and CD14+ monocytes occurred when AMD3100 was combined with G-CSF (Figure). In those receiving both AMD3100 and G-CSF, a fall in the % of T-cells expressing CCR7 and CXCR4 occurred 12 hours after the administration of AMD3100 compared to PB collected after 4 days of G-CSF; no other differences in the expression of a variety activation and/or adhesion molecules on T-cell subsets were observed. Whether differences in lymphocyte subsets mobilized with AMD3100 alone or in combination with G-CSF will impact immune reconstitution or other either immune sequela (i.e. GVHD, graft-vs-tumor) associated with allogeneic HCT is currently being assessed in an animal model of allogeneic transplantation.

Ex Vivo Fludarabine Selectively Depletes Human Naive T-Cell Subsets: A Step Toward Modifying Donor Lymphocyte Infusion.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1071-1071
Author(s):  
Melody M. Smith ◽  
Cynthia R. Giver ◽  
Edmund K. Waller ◽  
Christopher R. Flowers
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Memory T Cells ◽  
Ex Vivo ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Naïve T Cell ◽  
Naive T Cell

Abstract Ex vivo modification of donor lymphocytes with purine analogs (mDL) may help to minimize graft versus host disease (GvHD) while providing beneficial graft versus leukemia (GvL) effects. In a murine model system, we have shown that allogeneic donor splenocytes, treated with fludarabine ex vivo have significantly reduced GvHD activity when transferred to irradiated recipient mice, and retain anti-viral and GvL activities (Giver, 2003). This effect appears to be mediated by relative depletion of donor CD4 CD44low, “naive” T-cells. As a first step toward developing mDL for use in patients, we sought to evaluate the effects of ex vivo fludarabine exposure on human T-cell subsets, and to determine the minimum dose of fludarabine required to achieve this effect. Methods: Peripheral blood mononuclear cell samples from 6 healthy volunteers were evaluated at 0, 24, 48, and 72 hour time points after ex vivo incubation in varying dosages of fludarabine: 2, 5, and 10(n=3) mcg/ml. Fludarabine incubated samples were compared to samples that received no fludarabine (untreated). The total viable cell number was determined and the fractions and absolute numbers of viable CD4 and CD8 naïve and memory T-cells were determined using flow cytometry after incubation with 7-AAD (dead cell stain), CD4, CD8, CD45RA, CD62L, and CCR7 antibodies, and measuring the total viable cells/ml. Results: The numbers of viable CD4 and CD8 T-cells remained relatively stable in control cultures. Without fludarabine, the average viability at 72 hr of naive and memory T-cells were 92% and 77% for CD4 and 86% and 63% for CD 8 (Fig. 1A). Naive CD4 T-cells were more sensitive to fludarabine-induced death than memory CD4 cells. At 72 hr, the average viability of fludarabine-treated naive CD4 T-cells was 33% at 2 mcg/ml (8.2X the reduction observed in untreated cells) and 30% at 5 mcg/ml, while memory CD4 T-cells averaged 47% viability at 2 mcg/ml (2.3X the reduction observed in untreated cells) (Fig. 1B) and 38% at 5 mcg/ml. The average viability of naive CD8 T-cells at 72 hr was 27% at 2 mcg/ml and 20% at 5 mcg/ml, while memory CD8 T-cell viability was 22% at 2 mcg/ml and 17% at 5 mcg/ml. Analyses on central memory, effector memory, and Temra T-cells, and B-cell and dendritic cell subsets are ongoing. The 5 and 10 mcg/ml doses also yielded similar results in 3 initial subjects, suggesting that 2 mcg/ml or a lower dose of fludarabine is sufficient to achieve relative depletion of the naive T-cell subset. Conclusions: Future work will determine the minimal dose of fludarabine to achieve this effect, test the feasibility of using ex vivo nucleoside analog incubation to reduce alloreactivity in samples from patient/donor pairs, and determine the maximum tolerated dose of mDL in a phase 1 clinical trial with patients at high risk for relapse and infectious complications following allogeneic transplantation. Figure Figure

CD4 T lymphocytes are primed to express Fas ligand by CD4 cross-linking and to contribute to CD8 T-cell apoptosis via Fas/FasL death signaling pathway

Blood ◽  
2000 ◽  
Vol 96 (1) ◽  
pp. 195-202 ◽  
Author(s):  
Masaki Tateyama ◽  
Naoki Oyaizu ◽  
Thomas W. McCloskey ◽  
Soe Than ◽  
Savita Pahwa
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Interleukin 2 ◽  
Cd8 T Cell ◽  
T Cell Subsets ◽  
Cross Linking ◽  
Induced Apoptosis ◽  
Hiv 1

CD4 molecules serve as coreceptors for the T-cell receptor (TCR)/CD3 complex that are engaged coordinately with TCR and facilitate antigen-specific T-cell activation leading to interleukin 2 (IL-2) production and proliferation. However, cross-ligation of CD4 molecules prior to TCR stimulation has been shown to prime CD4 T cells to undergo apoptosis. Although in vivo and in vitro experiments have implicated the involvement of Fas/FasL interaction in this CD4 cross-linking (CD4XL)-induced apoptosis, detailed mechanisms to account for cell death induction have not been elucidated. In the present study, we demonstrate that CD4XL in purified T cells not only led to Fas up-regulation but also primed CD4 T cells to express FasL upon CD3 stimulation and rendered the T cells susceptible to Fas-mediated apoptosis. Notably, in addition to CD4+ T cells, CD4XL-induced sensitization for apoptosis was observed in CD8+ T cells as well and was associated with Bcl-x down-modulation. Both CD4 and CD8 T-cell subsets underwent apoptosis following cell–cell contact with FasL+ CD4 T cells. CD28 costimulation abrogated CD4XL/CD3-induced apoptosis with restoration of IL-2 production and prevented Bcl-x down-modulation. As CD4 molecules are the primary receptors for human immunodeficiency virus 1 (HIV-1), we conclude that HIV-1 envelope mediated CD4XL can lead to the generation of FasL-expressing CD4+ T cells that can lead to apoptosis of CD4 as well as CD8 T cells. These findings implicate a novel mechanism for CD8 T-cell depletion in HIV disease.

Sign in / Sign up

Export Citation Format

Share Document