Increased Expression of PD-1 on CD4+ T Cells from Patients with Chronic Lymphocytic Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4154-4154
Author(s):  
Mary M Sartor ◽  
David J Gottlieb
Keyword(s):  
T Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Nk Cells ◽  
Cd4 T Cells ◽  
Mononuclear Cells ◽  
Lymphocytic Leukemia ◽  
Effector Memory ◽  
Cd4 Cells ◽  
Normal Peripheral Blood

Abstract Although the predominant finding in patients with chronic lymphocytic leukemia (CLL) is an expansion of monoclonal B lymphocytes, a polyclonal expansion of T cells co-exists in CLL patients. Allogenic stem cell transplants for CLL suggest that a significant graft versus leukaemia effect mediated through recognition of minor MHC or leukaemia specific antigens can be achieved. Since it appears that the immune system and probably T cells recognise CLL cells, it is possible that one or more T cell defects might contribute to the initiation or maintenance of a clone of CLL lymphocytes. PD-1 is a coinhibitory molecule that is expressed on T cells in patients with chronic viral infections. It has been suggested that PD-1 expression might be a marker of cell exhaustion due to antigenic overstimulation. We examined the expression of PD-1 and its naturally occurring ligands PD-L1 and PD-L2 on both B and T cells in patients with CLL and compared this with expression on normal peripheral blood mononuclear cells. We found that PD-1 was expressed on over 10% of CD4+ T cells in 7 of 9 cases of CLL (mean 22±16%) but not on CD4+ T cells in any of 9 normal donors (mean 0±0%), p=0.0009. There was no difference in PD-1 expression on CD8+ or CD14+ PBMCs from CLL patients and normal donors (for CD8+ 24±21% and 19±16% for CLL and normals; for CD14+ 58±16% and 71±31% for CLL and normals). More than 10% of CD5+/19+ CLL cells expressed PD-1 in 7 of 10 cases (mean 18±18%) while more than 10% of normal B cells from 6 of 7 donors also expressed PD-1 (mean 49±30%). We examined the expression of PD-1 on naïve, central memory, effector memory and terminally differentiated subsets of CD4+ cells (CD62L+CD45RA+, CD62L+CD45RA−, CD62L−CD45RA− and CD62L−CD45RA+ respectively) from CLL patients and normal donors. The expression of PD-1 was higher on CD4+ cells from CLL patients in all subsets. The effect was most prominent in the effector memory subset (mean 54±4% for CLL patients versus 26±17% for normal donors, p=0.02). We looked for expression of PD-L1 and PD-L2 on T cells, B cells, monocytes and NK cells from CLL patients and normal donors. PD-L1 was only expressed on monocytes (mean 30±23%) and NK cells (mean 14±19%) from CLL patients and on monocytes from normal donors (mean 35±26%). There was no expression of PD-L2 on any cell type in either CLL patients or normal donors. We conclude that there is increased expression of the co-inhibitory molecule PD-1 on CD4+ T cells in patients with CLL. Ligation of PD-1 by PD-L1 expressed on monocytes or NK cells could inhibit immune responses to tumor and infectious antigens leading to persistence of clonally expanded cells and predisposition to opportunistic pathogens.

Anti-GvHD Effect of Antithymocyte Globulin Is Mediated by Antibodies Binding to T Cells and Regulatory NK Cells, and Less So or Not at All by Antibodies Binding to Other Mononuclear Cell Subsets

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2346-2346
Author(s):  
Mette Hoegh-Petersen ◽  
Minaa Amin ◽  
Yiping Liu ◽  
Alejandra Ugarte-Torres ◽  
Tyler S Williamson ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
B Cells ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Antithymocyte Globulin ◽  
Chronic Gvhd ◽  
Wilcoxon Test ◽  
Effector Memory

Abstract Abstract 2346 Introduction: Polyclonal rabbit-anti-human T cell globulin may decrease the likelihood of graft-vs-host disease (GVHD) without increasing the likelihood of relapse. We have recently shown that high levels of antithymocyte globulin (ATG) capable of binding to total lymphocytes are associated with a low likelihood of acute GVHD grade 2–4 (aGVHD) as well as chronic GVHD needing systemic therapy (cGVHD) but not increased likelihood of relapse (Podgorny PJ et al, BBMT 16:915, 2010). ATG is polyclonal, composed of antibodies for antigens expressed on multiple cell subsets, including T cells, B cells, NK cells, monocytes and dendritic cells. These cell subsets may play a role in the pathogenesis of GVHD. The anti-GVHD effect of ATG may be mediated through killing/inhibition of one or several of these cell subsets (eg, T cells) or their subsets (eg, naïve T cells as based on mouse experiments naïve T cells are thought to play a major role in the pathogenesis of GVHD). To better understand the mechanism of action of ATG on GVHD, we set out to determine levels of which ATG fraction (capable of binding to which cell subset) are associated with subsequent development of GVHD. Patients and Methods: A total of 121 patients were studied, whose myeloablative conditioning included 4.5 mg/kg ATG (Thymoglobulin). Serum was collected on day 7. Using flow cytometry, levels of the following ATG fractions were determined: capable of binding to 1. naïve B cells, 2. memory B cells, 3. naïve CD4 T cells, 4. central memory (CM) CD4 T cells, 5. effector memory (EM) CD4 T cells, 6. naïve CD8 T cells, 7. CM CD8 T cells, 8. EM CD8 T cells not expressing CD45RA (EMRA-), 9. EM CD8 T cells expressing CD45RA (EMRA+), 10. cytolytic (CD16+CD56+) NK cells, 11. regulatory (CD16-CD56high) NK cells, 12. CD16+CD56− NK cells, 13. monocytes and 14. dendritic cells/dendritic cell precursors (DCs). For each ATG fraction, levels in patients with versus without aGVHD or cGVHD were compared using Mann-Whitney-Wilcoxon test. For each fraction for which the levels appeared to be significantly different (p<0.05), we determined whether patients with high fraction level had a significantly lower likelihood of aGVHD or cGVHD than patients with low fraction level (high/low cutoff level was determined from ROC curve, using the point with maximum sum of sensitivity and specificity). This was done using log-binomial regression models, ie, multivariate analysis adjusting for recipient age (continuous), stem cell source (marrow or cord blood versus blood stem cells), donor type (HLA-matched sibling versus other), donor/recipient sex (M/M versus other) and days of follow up (continuous). Results: In univariate analyses, patients developing aGVHD had significantly lower levels of the following ATG fractions: binding to naïve CD4 T cells, EM CD4 T cells, naïve CD8 T cells and regulatory NK cells. Patients developing cGVHD had significantly lower levels of the following ATG fractions: capable of binding to naïve CD4 T cells, CM CD4 T cells, EM CD4 T cells, naïve CD8 T cells and regulatory NK cells. Patients who did vs did not develop relapse had similar levels of all ATG fractions. In multivariate analyses, high levels of the following ATG fractions were significantly associated with a low likelihood of aGVHD: capable of binding to naïve CD4 T cells (relative risk=.33, p=.001), EM CD4 T cells (RR=.30, p<.001), naïve CD8 T cells (RR=.33, p=.002) and regulatory NK cells (RR=.36, p=.001). High levels of the following ATG fractions were significantly associated with a low likelihood of cGVHD: capable of binding to naïve CD4 T cells (RR=.59, p=.028), CM CD4 T cells (RR=.49, p=.009), EM CD4 T cells (RR=.51, p=.006), naïve CD8 T cells (RR=.46, p=.005) and regulatory NK cells (RR=.55, p=.036). Conclusion: For both aGVHD and cGVHD, the anti-GVHD effect with relapse-neutral effect of ATG appears to be mediated by antibodies to antigens expressed on naïve T cells (both CD4 and CD8), EM CD4 T cells and regulatory NK cells, and to a lesser degree or not at all by antibodies binding to antigens expressed on B cells, cytolytic NK cells, monocytes or DCs. This is the first step towards identifying the antibody(ies) within ATG important for the anti-GVHD effect without impacting relapse. If such antibody(ies) is (are) found in the future, it should be explored whether such antibody(ies) alone or ATG enriched for such antibody(ies) could further decrease GVHD without impacting relapse. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Immunoglobulin secretory function of B cells from untreated patients with chronic lymphocytic leukemia and hypogammaglobulinemia: role of T cells

Blood ◽  
1983 ◽  
Vol 62 (4) ◽  
pp. 767-774 ◽  
Author(s):  
LA Fernandez ◽  
JM MacSween ◽  
GR Langley
Keyword(s):  
T Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
B Lymphocytes ◽  
Peripheral Blood ◽  
Cell Function ◽  
Lymphocytic Leukemia ◽  
Suppressor Activity ◽  
Normal Peripheral Blood

Abstract The mechanism of the hypogammaglobulinemia in patients with chronic lymphocytic leukemia (CLL) was studied by determining the generation of specific immunoglobulin-secreting cells in response to mitogen and antigen stimulation in culture. Normal peripheral blood B lymphocytes from 18 normal subjects cocultured with equal numbers of autologous T cells generated cells secreting 2,542 +/- 695 IgG, 2,153 +/- 615 IgA, and 2,918 +/- 945 IgM. Normal B lymphocytes cocultured with normal allogeneic T cells generated similar numbers. However, B lymphocytes from patients with chronic lymphocytic leukemia cocultured with T cells from the same patient generated only 0.5% as many cells secreting IgG and 11% and 23% as many secreting IgA and IgM, respectively. The reason for this markedly defective generation of immunoglobulin-secreting cells was investigated by evaluating T-helper, T-suppressor, and B-cell function using B cells from tonsil and T and B cells from peripheral blood of normal and leukemic individuals. T cells from patients with chronic lymphocytic leukemia provided somewhat greater help than did normal T cells to normal peripheral blood B cells and normal help to tonsil B cells, whether stimulated with mitogen or antigen. T cells from patients with chronic lymphocytic leukemia did not demonstrate increased suppressor function compared to normals with B cells from normal peripheral blood. The hypogammaglobulinemia in these patients therefore was associated with a markedly defective generation of immunoglobulin secreting cells, and as there was normal or increased T- cell helper activity without excessive suppressor activity, it seems likely that this was due to an intrinsic B-cell defect.

The effect of alemtuzumab maintenance therapy on B and T lymphocytes in chronic lymphocytic leukemia

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 7092-7092
Author(s):  
M. S. Kaufman ◽  
N. Driscoll ◽  
C. Johnson ◽  
A. Caramanica ◽  
D. Janson ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
T Lymphocytes ◽  
Maintenance Therapy ◽  
Cd4 T Cells ◽  
Opportunistic Infections ◽  
Lymphocytic Leukemia ◽  
Blood Levels ◽  
Time Point

7092 Background: We are conducting a pilot, exploratory study of the potential value of alemtuzumab(alem) in maintenance therapy of previously treated chronic lymphocytic leukemia (CLL) patients(pts) after they have achieved stable disease or partial remission with chemo or chemo-immunotherapy. We present the results of serially monitored CD19+ (B)lymphocytes and CD4+ (T) lymphocytes on eight evaluable patients. Methods: 30mg doses of alem were administered SC to all patients at the following schedule: wkly for 8 doses (8 wks), followed by q2 wks for 8 doses(16 wks), followed by q3 wks for 8 doses (24 wks). This schedule provides a total of 48 wks of maintenance treatment with alem. Patients received standard prophylaxis with sulfamethoxizole and acyclovir with regular CMV monitoring by quantitative PCR. Results: In the table we present data on the pattern of decrease in blood CD19+(B) cells and CD4+ (T) cells on eight evaluable pts at different time points after starting alem maintenance. Because flow cytometry was not done on all pts at each time point, the number of pts contributing to the calculation of mean counts at each given time point is variable. CD19+(B) cells were markedly reduced to 37% of baseline consistently, from 8 wks onward. CD4+(T) cells, on the other hand, were consistently higher than 50% of the baseline after 8 wks. No opportunistic infections were seen in any pt and treatment was well tolerated. Conclusion: These results from a single institution based pilot study demonstrate that alem used in maintenance schedule is effective in keeping the blood levels of CD19+(B) cells extremely low without concordant suppression of CD4+(T) lymphocytes. No significant financial relationships to disclose. [Table: see text]

scholarly journals Lenalidomide Induces Immunomodulation in Chronic Lymphocytic Leukemia and Enhances Antitumor Immune Responses Mediated by NK and CD4 T Cells

2014 ◽  
Vol 2014 ◽  
pp. 1-11 ◽  
Author(s):  
Andrea Acebes-Huerta ◽  
Leticia Huergo-Zapico ◽  
Ana Pilar Gonzalez-Rodriguez ◽  
Azahara Fernandez-Guizan ◽  
Angel R. Payer ◽  
...  
Keyword(s):  
T Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Nk Cells ◽  
Cd4 T Cells ◽  
Lymphocytic Leukemia ◽  
Leukemia Cells ◽  
Therapeutic Activity ◽  
Healthy Donors ◽  
Cd20 Expression ◽  
Dependent Cell

Lenalidomide is an immunomodulatory drug with therapeutic activity in chronic lymphocytic leukemia (CLL). However, it has pleiotropic effects, and the mechanism of action responsible for its therapeutic activity has not been well defined yet. Herein, we show that lenalidomide treatment does not have an effect on the proliferation of leukemia cells, but it increases the proliferation of B cells from healthy donors. Lenalidomide did not exert a direct effect on the apoptosis of leukemia cells obtained from CLL patients, although it indirectly induced their apoptosis through the activation of nonmalignant immune cells. Thus, lenalidomide markedly increased the proliferation of NK and CD4 T cells. The effect of lenalidomide on NK cells was secondary to the induction of IL-2 production by CD4 T cells. Accordingly, depletion of T cells or blockade of IL-2 activity completely abrogated the proliferation of NK cells. Additionally, lenalidomide enhanced NK and NKT-like cell-mediated natural cytotoxicity against leukemia cells from CLL patients. Lenalidomide also upregulated CD20 expression on leukemia cells and, accordingly, it had a synergistic effect with rituximab on promoting antibody-dependent cell-mediated cytotoxicity against primary leukemia cells. Overall, these observations provide a support for combining lenalidomide with rituximab as a treatment in CLL.

scholarly journals Broad phenotypic alterations and potential dysfunction of lymphocytes in individuals clinically recovered from COVID-19

2021 ◽  
Author(s):  
Jingyi Yang ◽  
Maohua Zhong ◽  
Ejuan Zhang ◽  
Ke Hong ◽  
Qingyu Yang ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Cd4 T Cells ◽  
Mononuclear Cells ◽  
Cell Subset ◽  
Effector Memory ◽  
Peripheral Blood Mononuclear ◽  
Follicular Helper ◽  
Hla Dr ◽  
Ifn Γ

Abstract Although millions of patients have clinically recovered from COVID-19, little is known about the immune status of lymphocytes in these individuals. In this study, the peripheral blood mononuclear cells (PBMCs) of a clinically recovered (CR) cohort were comparatively analyzed with those of an age- and sex-matched healthy donor (HD) cohort. We found that CD8+ T cells in the CR cohort had higher numbers of effector T cells and effector memory T cells but lower Tc1 (IFN-γ+), Tc2 (IL-4+), and Tc17 (IL-17A+) cell frequencies. The CD4+ T cells of the CR cohort were decreased in frequency, especially the central memory T cell subset. Moreover, CD4+ T cells in the CR cohort showed lower PD-1 expression and had lower frequencies of Th1 (IFN-γ+), Th2 (IL-4+), Th17 (IL-17A+), and circulating follicular helper T (CXCR5+PD-1+) cells. Accordingly, the proportion of isotype-switched memory B cells (IgM−CD20hi) among B cells in the CR cohort showed a significantly lower proportion, although the level of the activation marker CD71 was elevated. For CD3−HLA-DR− lymphocytes in the CR cohort, in addition to lower levels of IFN-γ, granzyme B, and T-bet, the correlation between T-bet and IFN-γ was not observed. Additionally, by taking into account the number of days after discharge, all the phenotypes associated with reduced function did not show a tendency toward recovery within 4‒11 weeks. The remarkable phenotypic alterations in lymphocytes in the CR cohort suggest that SARS-CoV-2 infection profoundly affects lymphocytes and potentially results in dysfunction even after clinical recovery.

scholarly journals MiR-181b in Chronic Lymphocytic Leukemia B Cells Is Regulated By Cellular Interaction with CD4+ T Cells and Increases the CTL Toxicity Versus the Leukemic Clone

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4134-4134
Author(s):  
Mirco di Marco ◽  
Serena Veschi ◽  
Rosa Visone ◽  
Giuseppe Leone ◽  
Paola Lanuti ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Cd4 T Cells ◽  
Lymphocytic Leukemia ◽  
Healthy Donors ◽  
Leukemic Clone

Abstract Clinical progression of chronic lymphocytic leukemia (CLL) is characterized by gradual reduction of the ratio T/B cells, along with immune cell dysfunction due, at least in part, to T cell defects, such as decreased expression of CD40L and reduced signaling via the TCR CD3. This compromise the ability of T cells to respond and to eliminate leukemic cell from CLL patients. Enhanced activation of either allogenic or autologous T cells can drive the death of CLL cells in vitro and in human subjects. Changes in microRNAs expression also characterize clinical progression of CLL with a strong decrease of miR-181b/a and miR-130a associated with the more aggressive phase of the disease. The miR-181b targets anti-apoptotic proteins, such as BCL-2 and MCL1 and its expression correlates with those protein levels in CLL. In this study we demonstrate that the expression of those microRNAs in CLL-B cells, are regulated by T cells. We co-cultured allogenic pure CLL-B cells with either activated (CD2, CD3 and CD28 antibodies, used to mimic antigen-presenting cells) or not activated CD4+ T cells from healthy donors. We observed a significant increase of mir-181b/a and miR-130a expression in CLL B-cells after co-culture with activated CD4+ T cells in 8 out of 11 cases. A significant increase of these miRs was also determined in purified CLL B-cells after 4 days activation of peripheral blood mononuclear cells (PBMCs) from CLL patients, even if in minor rate. By the use of specific antibodies, co-culture with Hela CD40 expressing cells and transwell experiments, we established that this effect is a T/B contact-dependent signaling mediated through CD40L-CD40 interaction. We determine that increased expression of the 3 miRs occurs at the transcriptional level. Since the expression of miR-181b showed the most significant variation in previous experiments it was selected for further analyses. We next investigated the in vivo role of the miR-181b in highly immunodeficient mice. The CLL cell line, MEC-01, infected with either the LV-miR-181b_coGFP or the LV-CTRL_coGFP was intravenously inoculated in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice. Mice were sacrificed after 4 weeks and assayed for percentage of GFP+ cells in bone marrow and spleen compartments. The miR-181b did not show any specific effect into the leukemic clone. However when the same cells were inoculated in an environment hosting mature T cells, miR-181b consistently influences the death of leukemic cells (Fig 1B), suggesting that T cells are required to potentiate the apoptotic role of this miRNA. To explain what we observed in vivo, we mixed in vitro MEC-01 infected with either the LV-miR-181b or the LV-CTRL and CD8+ T cells from healthy donors. After few hours of contact T cells showed stronger cytotoxic effect on MEC-01 carrying miR-181b as compared to the control. Mixed lymphocyte reaction CD40L-activated CLL and T cells is used to generate effector CTLs. Therefore we grew T cell with CD40L-activated MEC-01 in which the expression of miR-181b was either shut down by lentiviral vector or unchanged as control. After one week, we monitored by cytofluorimetry the CD38 surface marker on T cells since its expression has been associated with more active CTLs and, by ELISA, the release of IL-10, the inhibitor of the potent inducer of CTLs INF-g. We demonstrate that activated MEC-01 with higher expression of miR-181b leads to an increase of the cell number expressing CD38 and this was accompanied by a reduced release of IL-10 from B cells through down-regulation of c-FOS, which we show to be target of the miR-181b and to promote the transcription of the IL-10. In conclusion, our data suggest a role of the miR-181b in the immune response against CLL-B cells. We show that an efficient activation of CD4+ T cells through CD3-complex pathway and a right CD40L-CD40 interaction lead to a significant increase of the some miRNAs deregulated over the progression of chronic lymphocytic leukemia, namely miR-181b. This miRNA potentiates the cytotoxicity of T cells favoring the killing of the leukemic clone. Disclosures No relevant conflicts of interest to declare.

Induction of Alloantigen Specific CD4 T Cell Anergy by B7: CD28 Mediated Costimulatory Blockade Significantly Alters the Functional Program of Bystander CD8, Monocytes, NK and B Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3656-3656
Author(s):  
Gullu Gorgun ◽  
Fenglong Liu ◽  
Eleanor Howe ◽  
Patrick Philpot ◽  
Eva Guinan ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Nk Cells ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Mononuclear Cells ◽  
Global Gene Expression ◽  
Differentially Expressed ◽  
Functional Program ◽  
The Impact

Abstract Retention of antigen (Ag) specific immunity to pathogens and tumor cells in the context of adequate control of alloreactivity remains a challenge for allogeneic hematopoietic stem cell transplantation (HSCT). Global or subset depletion of T cells achieves control of alloreactivity at the cost of loss of non-allospecific repertoire. Induction of Ag specific anergy has been explored as a way to selectively impact alloreactivity. Anergy is induced when Ag is presented to CD4 T cells without CD28:B7 mediated costimulation. We and others have shown that CD4 anergy results from a positive signaling cascade, rendering Ag specific T cells unable to proliferate and produce cytokines on Ag specific rechallenge. When this concept was translated to a haploidentical HSCT clinical trial, all donor marrow harvest mononuclear cells were subjected to inhibition of CD28 mediated costimulation by CTLA-4-Ig, a fusion protein binding both B7.1 and B7.2 costimulatory molecules. In our ongoing clinical trial, unfractionated donor peripheral blood mononuclear cells (PBMC) are cultured with allostimulators in the presence of costimulatory blockade (CSB) provided by anti-B7.1 and anti-B7.2 humanized monoclonal antibodies. To understand the impact of inducing alloAg specific CD4 T cell anergy on bystander PBMC, we investigated the effects on CD8 T cells, monocytes, B and NK cells. Mimicking our ex vivo clinical anergization protocol, primary MLR using PBMC from fully HLA mismatched healthy donors (n=12) were performed in the presence or absence of CSB. CSB resulted in 73% mean inhibition of proliferation after 72 hrs of primary MLR. CD4 and CD8 T cells, monocytes, NK and B cells from these MLRs were isolated by negative selection as were control unmanipulated CD4 and CD8 T cells. From each population, RNA was extracted and global gene expression profiling performed using Affymetrix human 133plus2 chips. Unsupervised analysis was performed using DNA-Chip Analyzer and supervised analysis using Significance Analysis for Microarrays. While the frequency of alloreactive CD4 T cells in human PBMC is only 1–10%, we observed global gene expression variance (436 genes) between unstimulated CD4 cells vs. CD4 cells isolated from MLR in the presence or absence of CSB (P≤0.05). Even more surprising was the impact of CSB on bystander CD8, monocytes, NK and B cells. Between cells from MLR with or without CSB, there were 632 differentially expressed genes in CD8 T cells, 105 differentially expressed in NK cells, 85 differentially expressed in monocytes and 1781 in B cells (P≤0.05 for all populations tested). We observed not only expected alterations (e.g, in inflammatory cytokines and receptor mediated signaling) but also observed changes [e.g. in proteasome degradation (i.e. CD4, CD8, NK and B cells) and in expression of genes regulating cell motility (i.e. CD8 T and NK cells)]. These results demonstrate that induction of alloAg specific anergy within a small population of alloreactive CD4 T cells results in dramatic alterations of the genetic repertoire of bystander cells. Thus, blockade of the CD28:B7 costimulatory pathway not only inactivates alloreactive CD4 T cells but also alters the functional program of other immune cells that will repopulate the host post-HSCT.

scholarly journals Pharmacologic Inhibition with Duvelisib (IPI-145) and Genetic Inhibition of PI3K p110δ Antagonizes Intrinsic and Extrinsic Survival Signals in Chronic Lymphocytic Leukemia (CLL)

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3324-3324 ◽  
Author(s):  
Shuai Dong ◽  
Daphne Guinn ◽  
Jason A Dubovsky ◽  
Yiming Zhong ◽  
Amy Lehman ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Nk Cells ◽  
Cell Line ◽  
Research Funding ◽  
Lymphocytic Leukemia ◽  
Pi3k Signaling

Abstract Background: Chronic lymphocytic leukemia (CLL), the most common adult leukemia in the western world, remains an incurable B-cell malignancy. It is characterized by the accumulation of malignant mature B cells in the blood, lymph nodes, spleen, and bone marrow. CLL cells display up-regulated B-cell receptor (BCR) activation, which maintains B cell survival and proliferation through transmitting microenvironmental stimuli. Due to aberrant regulation of the BCR, CLL cells display constitutively activated survival and proliferation pathways, such as phosphoinositide-3 kinase (PI3K) and Bruton’s tyrosine kinase (BTK) pathways. Small molecules that target such kinases in the BCR pathway have shown significant clinical activity in CLL patients. Both the PI3K p110δ inhibitor, idelalisib, and the BTK inhibitor, ibrutinib, have received approval by FDA for treatment of relapsed CLL. However, patients still relapse on these therapies. Methods/Results: Here we use pharmacologic and genetic approaches to further characterize the role of PI3K signaling in the leukemia pathogenesis in the CLL cell and in the microenvironment. We describe that a PI3K p110δ and p110γ inhibitor, duvelisib (IPI-145), which is in late stage clinical development, attenuates pro-survival signals in the OSU-CLL cell line and primary human and murine CLL cells and promotes apoptosis and downstream pathway inactivation in primary human and murine CLL cells in a dose- and time-dependent fashion. To examine the cytotoxicity of duvelisib in normal immune cells, we incubated whole blood from CLL patients with 0.25-5 μM duvelisib for 48 hours and analyzed by flow cytometry for absolute count of live CD3+ T cells, CD56+ NK cells and CD19+ B cells. T cells and NK cells were sensitive to duvelisib, displaying about 20% decrease in viability at concentrations greater than 0.5μM; however, the B cell population showed about 50% decrease in viability. To specifically examine normal B cells, we isolated CD19+ B cells from healthy volunteer blood and incubated with 1 μM duvelisib for 48 hours and observed no cytotoxicity, despite observing a significant decrease in CLL cells viability under the same conditions. Additionally, duvelisib is highly effective at reducing downstream PI3K signaling in a B cell line with the ibrutinib resistance conferring BTK C481S mutation. Genetically we show that the PI3K p110δ-inactivating and the TCL1 leukemia murine models can be utilized to further explore the differential role of PI3K p110δ in the leukemic cell and microenvironment. Our study indicates that systemic disruption of PI3K p110δ function in the TCL1 mouse significantly prevents spontaneous leukemia development, indicating that PI3K p110δ is a critical kinase for CLL disease initiation and expansion. Moreover, inactivation of PI3K p110δ in the microenvironment showed a dose-dependent effect in delaying leukemia engraftment. This suggests that PI3K p110δ activity is also critical in the non-B cell compartment for leukemia progression. While our group has focused on the role of PI3K p110δ, we continue to examine the role of PI3K p110γ using the PI3K p110δ and p110γ inhibitor, duvelisib. Disclosures Dubovsky: Principia Inc.: Research Funding. Kutok:Infinity Pharmaceuticals, Inc.: Employment, Equity Ownership. Byrd:Pharmacyclics: Research Funding.

scholarly journals Relation of Neutrophil Gelatinase-Associated Lipocalin Overexpression to the Resistance to Apoptosis of Tumor B Cells in Chronic Lymphocytic Leukemia

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2124 ◽  
Author(s):  
Brigitte Bauvois ◽  
Elodie Pramil ◽  
Ludovic Jondreville ◽  
Elise Chapiro ◽  
Claire Quiney ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Mononuclear Cells ◽  
Critical Role ◽  
Leukemia Cell ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Normal Peripheral Blood ◽  
Neutrophil Gelatinase Associated Lipocalin ◽  
Neutrophil Gelatinase

The resistance to apoptosis of chronic lymphocytic leukemia (CLL) cells partly results from the deregulated production of survival signals from leukemic cells. Despite the development of new therapies in CLL, drug resistance and disease relapse still occur. Recently, neutrophil gelatinase-associated lipocalin (NGAL), a secreted glycoprotein, has been suggested to have a critical role in the biology of tumors. Thus, we investigated the relevance of NGAL in CLL pathogenesis, analyzed the expression of its cellular receptor (NGAL-R) on malignant B cells and tested whether CLL cells are resistant to apoptosis through an autocrine process involving NGAL and NGAL-R. We observed that NGAL concentrations were elevated in the serum of CLL patients at diagnosis. After treatment (and regardless of the therapeutic regimen), serum NGAL levels normalized in CLL patients in remission but not in relapsed patients. In parallel, NGAL and NGAL-R were upregulated in leukemic cells from untreated CLL patients when compared to normal peripheral blood mononuclear cells (PBMCs), and returned to basal levels in PBMCs from patients in remission. Cultured CLL cells released endogenous NGAL. Anti-NGAL-R antibodies enhanced NGAL-R+ leukemia cell death. Conversely, recombinant NGAL protected NGAL-R+ CLL cells against apoptosis by activating a STAT3/Mcl-1 signaling pathway. Our results suggest that NGAL and NGAL-R, overexpressed in untreated CLL, participate in the deregulation of the apoptotic machinery in CLL cells, and may be potential therapeutic clues for CLL treatment.

scholarly journals Natural Killer Cell Dysfunction in Chronic Lymphocytic Leukemia Is Associated with Loss of the Mature KIR3DL1+ Subset

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3318-3318 ◽  
Author(s):  
Alexander W. MacFarlane ◽  
Mowafaq Jillab ◽  
Mitchell R Smith ◽  
R. Katherine Alpaugh ◽  
Marion E. Cole ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Nk Cells ◽  
Peripheral Blood ◽  
Cell Function ◽  
Nk Cell ◽  
Tumor Burden ◽  
Lymphocytic Leukemia ◽  
Healthy Controls

Abstract Background: B-cell chronic lymphocytic leukemia (CLL) is a common blood cancer characterized by high prevalence of malignant B cells in peripheral blood. Small lymphocytic lymphoma (SLL) is considered to be a different presentation of the same disease, with the malignant B cells primarily localized in lymph nodes. Natural killer (NK) cells are innate immune effectors that can spontaneously identify and kill malignant cells, especially hematopoietic cancers. In peripheral blood of CLL patients, NK cells are chronically exposed to significant tumor burden, which is predicted to influence their phenotype and function. Effective NK cell function may be particularly beneficial in CLL patients, since commonly-used monoclonal antibody therapies (e.g. rituximab, alemtuzumab) rely at least partially on ADCC-mediated by NK cells. Methods: We performed a prospective analysis of biomarkers on fresh peripheral blood lymphocytes from 25 untreated CLL patients, 10 untreated SLL and 17 age-matched healthy controls by 10-color flow cytometry. All subjects signed IRB approved informed consent forms. Our study analyzed 180 distinct biomarker parameters, with a particular focus on NK and T cells. Differences in biomarker expression between patients with SLL, CLL, and healthy controls were compared by Wilcoxon rank-sum test. Results: Absolute numbers of NK and T cells per µl of blood were significantly higher in CLL patients, and this correlated with increased B cell numbers. As indicators of immune suppression, the frequency of regulatory T cells was significantly increased in CLL samples, as were levels of PD-1 expression on T cells and CD56dim NK cells. NK cells in CLL expressed higher levels of CD27, which is characteristic of a less mature phenotype, and CD56dim cells expressed lower levels of NKG2D. Compared to healthy controls, CLL samples displayed a marked reduction in degranulation by CD56dim NK cells in response to transformed 721.221 B cells, either with or without rituximab. CD56dim NK cells from CLL patients were also less viable under resting conditions or when challenged with target cells, especially in ADCC responses. We further observed a striking reduction in the frequency and viability of KIR3DL1+ NK cells, which progressed over time in most CLL patients. Surprisingly, CLL patients with the highest levels of PD-1 expression on NK cells possessed genes for both KIR3DL1 and its ligand, HLA-Bw4. Our findings were also clearly evident in a CLL patient compared to her healthy monozygotic twin, thereby providing compelling support for the results in the full patient cohort. The altered expression levels of nearly all of the NK cell biomarkers and degranulation were less pronounced in blood samples from SLL patients, presumably due to low tumor burden in peripheral blood. Conclusions: CLL patients have increased numbers of NK cells in peripheral blood, but these NK cells are less mature, are significantly depleted of the KIR3DL1+ subset, and have deficits in degranulation response, reduced expression of NKG2D activating receptor, increased expression of inhibitory PD-1, and enhanced susceptibility to activation-induced death when challenged with tumor targets and rituxumab. Our findings support the hypothesis that immune dysfunction in CLL may be due in part to a selective loss of mature KIR3DL1+ NK cells, possibly upon encountering overwhelming tumor burden in peripheral blood, and CLL patients may benefit from therapeutic strategies that augment NK cell function. Disclosures No relevant conflicts of interest to declare.

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