Reduced frequencies and suppressive function of CD4+CD25hi regulatory T cells in patients with chronic lymphocytic leukemia after therapy with fludarabine

Abstract Globally suppressed T-cell function has been described in many patients with cancer to be a major hurdle for the development of clinically efficient cancer immunotherapy. Inhibition of antitumor immune responses has been mainly linked to inhibitory factors present in cancer patients. More recently, increased frequencies of CD4+CD25hi regulatory T cells (Treg cells) have been described as an additional mechanism reducing immunity. We assessed 73 patients with B-cell chronic lymphocytic leukemia (CLL) and 42 healthy controls and demonstrated significantly increased frequencies of cytotoxic T lymphocyte-associated protein 4 (CTLA4+)–, Forkhead box P3 (FOXP3+)–, glucocorticoid-induced tumor necrosis factor receptor-related protein (GITR+)–, CD62L+–, transforming growth factor β1 (TGF-β1+)–, interleukin 10 (IL-10+)–Treg cells in patients with CLL, with highest frequencies in untreated or progressing patients presenting with extended disease. Most surprisingly, in the majority of patients with CLL treated with fludarabine-containing therapy regimens the inhibitory function of Treg cells was decreased or even abrogated. In addition, frequencies of Treg cells were significantly decreased after therapy with fludarabine. In light of similar findings for cyclophosphamide the combination of fludarabine and cyclophosphamide might be further exploited in strategies reducing immunosuppression prior to cancer immunotherapy.

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Immunosuppressive regulatory T cells are abundant in the reactive lymphocytes of Hodgkin lymphoma

Blood ◽

10.1182/blood-2003-07-2594 ◽

2004 ◽

Vol 103 (5) ◽

pp. 1755-1762 ◽

Cited By ~ 361

Author(s):

Neil A. Marshall ◽

Linsey E. Christie ◽

Laura R. Munro ◽

Dominic J. Culligan ◽

Peter W. Johnston ◽

...

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Hodgkin Lymphoma ◽

Mononuclear Cells ◽

Cell Function ◽

Cytotoxic T Lymphocyte ◽

Interleukin 10 ◽

Cell Contact ◽

T Helper 1 ◽

Peripheral Blood Mononuclear

Abstract Although immunosuppression has long been recognized in Hodgkin lymphoma (HL), the underlying basis for the lack of an effective immune response against the tumor remains unclear. The aim was to test our hypothesis that regulatory T cells dominate involved lymph nodes. The approach was to assay CD4+ T-cell function in HL-infiltrating lymphocytes (HLILs) and paired peripheral blood mononuclear cells (PBMCs) of 24 patients. Strikingly, unlike PBMCs, HLILs were anergic to stimulation with mitogen, primary, or recall antigens, mounting no proliferative responses and only rare T-helper 1 (Th1) or Th2 cytokine responses. Mixing paired HLILs and PBMCs showed the anergic effect was dominant and suppressed PBMC responses. Furthermore, flow cytometry demonstrated that HLILs contained large populations of both interleukin-10 (IL-10)–secreting T-regulatory 1 (Tr1) and CD4+CD25+ regulatory T cells. We found evidence for 3 mechanisms of action implicated in the suppressive functions of regulatory T cells: the inhibition of PBMCs by HLILs was ameliorated by neutralizing IL-10, by preventing cell-to-cell contact, and by blocking anti–cytotoxic T lymphocyte–associated antigen 4 (anti–CTLA-4). Thus, HLILs are highly enriched for regulatory T cells, which induce a profoundly immunosuppressive environment and so provide an explanation for the ineffective immune clearance of Hodgkin-Reed Sternberg cells.

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Neuropilin-1highCD4+CD25+ Regulatory T Cells Exhibit Primary Negative Immunoregulation in Sepsis

Mediators of Inflammation ◽

10.1155/2016/7132158 ◽

2016 ◽

Vol 2016 ◽

pp. 1-11 ◽

Cited By ~ 12

Author(s):

Yu-Lei Gao ◽

Yan-Fen Chai ◽

An-Long Qi ◽

Ying Yao ◽

Yan-Cun Liu ◽

...

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Polyclonal Antibody ◽

Transforming Growth Factor ◽

Cytotoxic T Lymphocyte ◽

Cecal Ligation ◽

Interleukin 10 ◽

Neuropilin 1 ◽

The Impact

Regulatory T cells (Tregs) appear to be involved in sepsis-induced immune dysfunction; neuropilin-1 (Nrp-1) was identified as a surface marker for CD4+CD25+Tregs. In the current study, we investigated the negative immunoregulation of Nrp-1highCD4+CD25+Tregs and the potential therapeutic value of Nrp-1 in sepsis. Splenic CD4+CD25+Tregs from cecal ligation and puncture (CLP) mouse models were further segregated into Nrp-1highTregs and Nrp-1lowTregs; they were cocultured with CD4+CD25− T cells. The expression of forkhead/winged helix transcription factor-3 (Foxp-3), cytotoxic T-lymphocyte associated antigen-4 (CTLA-4), membrane associated transforming growth factor-β (TGF-βm+), apoptotic rate, and secretive ability [including TGF-β and interleukin-10 (IL-10)] for various types of Tregs, as well as the immunosuppressive ability of Tregs on CD4+CD25− T cells, were determined. Meanwhile, the impact of recombinant Nrp-1 polyclonal antibody on the demethylation of Foxp-3-TSDR (Treg-specific demethylated region) was measured in in vitro study. Sepsis per se markedly promoted the expression of Nrp-1 of CD4+CD25+Tregs. Foxp-3/CTLA-4/TGF-βm+ of Nrp-1highTregs were upregulated by septic challenge. Nrp-1highTregs showed strong resilience to apoptosis and secretive ability and the strongest immunosuppressive ability on CD4+CD25− T cells. In the presence of lipopolysaccharide (LPS), the recombinant Nrp-1 polyclonal antibody reduced the demethylation of Foxp-3-TSDR. Nrp-1highTregs might reveal primary negative immunoregulation in sepsis; Nrp-1 could represent a new potential therapeutic target for the study of immune regulation in sepsis.

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Interleukin-10 suppression enhances T-cell antitumor immunity and responses to checkpoint blockade in chronic lymphocytic leukemia

10.1101/2020.07.15.204560 ◽

2020 ◽

Author(s):

J.R. Rivas ◽

S.S. Alhakeem ◽

Y. Liu ◽

J.M. Eckenrode ◽

F. Marti ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Chronic Lymphocytic Leukemia ◽

Cell Function ◽

Cell Activity ◽

Interleukin 10 ◽

Interferon Γ ◽

Immune Checkpoint Blockade ◽

Lymphocytic Leukemia ◽

Checkpoint Blockade

AbstractT-cell dysfunction is a hallmark of B-cell Chronic Lymphocytic Leukemia (CLL). CLL cells downregulate T-cell responses by expressing regulatory molecules including programmed death ligand-1 (PD-L1) and Interleukin-10 (IL-10). Immune checkpoint blockade (ICB) aims to restore T-cell function by preventing the ligation of inhibitory receptors like PD-1, however most CLL patients do not respond well to this therapy. Thus, we investigated whether IL-10 suppression could enhance antitumor T-cell activity and improve responses to ICB. Since CLL IL-10 expression depends on Sp1, we utilized a novel, better tolerated mithramycin analogue, MTMox32E, to suppress CLL IL-10. We found MTMox32E inhibited mouse and human CLL IL-10 production and maintained T-cell effector function. In the Eμ-Tcl1 mouse model, treatment reduced plasma IL-10 and CLL burden while it increased CD8+ T-cell proliferation, effector and memory cell prevalence, and interferon-γ production. When combined with ICB, suppression of IL-10 improved responses to anti-PD-L1 as shown by a 4.5-fold decrease in CLL cell burden compared with anti-PD-L1 alone. Combination therapy also produced more interferon-γ+, cytotoxic effector KLRG1+, and memory CD8+ T-cells, with fewer exhausted T-cells than ICB alone. Since current therapies for CLL do not target IL-10, this provides a novel strategy to increase the efficacy of T-cell-based immunotherapies.

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Targeting CD38 is lethal to Breg-like chronic lymphocytic leukemia cells and Tregs, but restores CD8+ T-cell responses

Blood Advances ◽

10.1182/bloodadvances.2019001091 ◽

2020 ◽

Vol 4 (10) ◽

pp. 2143-2157 ◽

Cited By ~ 1

Author(s):

Alak Manna ◽

Timothy Kellett ◽

Sonikpreet Aulakh ◽

Laura J. Lewis-Tuffin ◽

Navnita Dutta ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Chronic Lymphocytic Leukemia ◽

Cd8 T Cells ◽

Mononuclear Cells ◽

Transforming Growth Factor ◽

Ex Vivo ◽

Xenograft Model ◽

Lymphocytic Leukemia ◽

Dependent Manner

Abstract Patients with chronic lymphocytic leukemia (CLL) are characterized by monoclonal expansion of CD5+CD23+CD27+CD19+κ/λ+ B lymphocytes and are clinically noted to have profound immune suppression. In these patients, it has been recently shown that a subset of B cells possesses regulatory functions and secretes high levels of interleukin 10 (IL-10). Our investigation identified that CLL cells with a CD19+CD24+CD38hi immunophenotype (B regulatory cell [Breg]–like CLL cells) produce high amounts of IL-10 and transforming growth factor β (TGF-β) and are capable of transforming naive T helper cells into CD4+CD25+FoxP3+ T regulatory cells (Tregs) in an IL-10/TGF-β-dependent manner. A strong correlation between the percentage of CD38+ CLL cells and Tregs was observed. CD38hi Tregs comprised more than 50% of Tregs in peripheral blood mononuclear cells (PBMCs) in patients with CLL. Anti-CD38 targeting agents resulted in lethality of both Breg-like CLL and Treg cells via apoptosis. Ex vivo, use of anti-CD38 monoclonal antibody (mAb) therapy was associated with a reduction in IL-10 and CLL patient-derived Tregs, but an increase in interferon-γ and proliferation of cytotoxic CD8+ T cells with an activated phenotype, which showed an improved ability to lyse patient-autologous CLL cells. Finally, effects of anti-CD38 mAb therapy were validated in a CLL–patient-derived xenograft model in vivo, which showed decreased percentage of Bregs, Tregs, and PD1+CD38hiCD8+ T cells, but increased Th17 and CD8+ T cells (vs vehicle). Altogether, our results demonstrate that targeting CD38 in CLL can modulate the tumor microenvironment; skewing T-cell populations from an immunosuppressive to immune-reactive milieu, thus promoting immune reconstitution for enhanced anti-CLL response.

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Regulatory T cells in chronic lymphocytic leukemia: implication for immunotherapeutic interventions

Tumor Biology ◽

10.1007/s13277-013-0832-x ◽

2013 ◽

Vol 34 (4) ◽

pp. 2031-2039 ◽

Cited By ~ 33

Author(s):

Farhad Jadidi-Niaragh ◽

Ghasem Ghalamfarsa ◽

Mehdi Yousefi ◽

Mina Hajifaraj Tabrizi ◽

Fazel Shokri

Keyword(s):

T Cells ◽

Chronic Lymphocytic Leukemia ◽

Regulatory T Cells ◽

Lymphocytic Leukemia

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Secretome screening reveals immunomodulating functions of IFNα-7, PAP and GDF-7 on regulatory T-cells

Scientific Reports ◽

10.1038/s41598-021-96184-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Mei Ding ◽

Rajneesh Malhotra ◽

Tomas Ottosson ◽

Magnus Lundqvist ◽

Aman Mebrahtu ◽

...

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Cell Function ◽

Foxp3 Expression ◽

Treg Cells ◽

Secreted Proteins ◽

External Stimulation ◽

Marker Proteins ◽

Positive Regulators ◽

Conformational Model

AbstractRegulatory T cells (Tregs) are the key cells regulating peripheral autoreactive T lymphocytes. Tregs exert their function by suppressing effector T cells. Tregs have been shown to play essential roles in the control of a variety of physiological and pathological immune responses. However, Tregs are unstable and can lose the expression of FOXP3 and suppressive functions as a consequence of outer stimuli. Available literature suggests that secreted proteins regulate Treg functional states, such as differentiation, proliferation and suppressive function. Identification of secreted proteins that affect Treg cell function are highly interesting for both therapeutic and diagnostic purposes in either hyperactive or immunosuppressed populations. Here, we report a phenotypic screening of a human secretome library in human Treg cells utilising a high throughput flow cytometry technology. Screening a library of 575 secreted proteins allowed us to identify proteins stabilising or destabilising the Treg phenotype as suggested by changes in expression of Treg marker proteins FOXP3 and/or CTLA4. Four proteins including GDF-7, IL-10, PAP and IFNα-7 were identified as positive regulators that increased FOXP3 and/or CTLA4 expression. PAP is a phosphatase. A catalytic-dead version of the protein did not induce an increase in FOXP3 expression. Ten interferon proteins were identified as negative regulators that reduced the expression of both CTLA4 and FOXP3, without affecting cell viability. A transcriptomics analysis supported the differential effect on Tregs of IFNα-7 versus other IFNα proteins, indicating differences in JAK/STAT signaling. A conformational model experiment confirmed a tenfold reduction in IFNAR-mediated ISG transcription for IFNα-7 compared to IFNα-10. This further strengthened the theory of a shift in downstream messaging upon external stimulation. As a summary, we have identified four positive regulators of FOXP3 and/or CTLA4 expression. Further exploration of these Treg modulators and their method of action has the potential to aid the discovery of novel therapies for both autoimmune and infectious diseases as well as for cancer.

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CD4+ regulatory T cells require CTLA-4 for the maintenance of systemic tolerance

Journal of Experimental Medicine ◽

10.1084/jem.20081811 ◽

2009 ◽

Vol 206 (2) ◽

pp. 421-434 ◽

Cited By ~ 160

Author(s):

Randall H. Friedline ◽

David S. Brown ◽

Hai Nguyen ◽

Hardy Kornfeld ◽

JinHee Lee ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Cell Function ◽

Cytotoxic T Lymphocyte ◽

Critical Role ◽

Lymphoproliferative Disease ◽

In Trans ◽

And Function

Cytotoxic T lymphocyte antigen-4 (CTLA-4) plays a critical role in negatively regulating T cell responses and has also been implicated in the development and function of natural FOXP3+ regulatory T cells. CTLA-4–deficient mice develop fatal, early onset lymphoproliferative disease. However, chimeric mice containing both CTLA-4–deficient and –sufficient bone marrow (BM)–derived cells do not develop disease, indicating that CTLA-4 can act in trans to maintain T cell self-tolerance. Using genetically mixed blastocyst and BM chimaeras as well as in vivo T cell transfer systems, we demonstrate that in vivo regulation of Ctla4−/− T cells in trans by CTLA-4–sufficient T cells is a reversible process that requires the persistent presence of FOXP3+ regulatory T cells with a diverse TCR repertoire. Based on gene expression studies, the regulatory T cells do not appear to act directly on T cells, suggesting they may instead modulate the stimulatory activities of antigen-presenting cells. These results demonstrate that CTLA-4 is absolutely required for FOXP3+ regulatory T cell function in vivo.

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Immunosuppressive Treg cells acquire the phenotype of effector-T cells in chronic lymphocytic leukemia patients

Journal of Translational Medicine ◽

10.1186/s12967-018-1545-0 ◽

2018 ◽

Vol 16 (1) ◽

Cited By ~ 7

Author(s):

Serena De Matteis ◽

Chiara Molinari ◽

Giulia Abbati ◽

Tania Rossi ◽

Roberta Napolitano ◽

...

Keyword(s):

T Cells ◽

Chronic Lymphocytic Leukemia ◽

Treg Cells ◽

Effector T Cells ◽

Lymphocytic Leukemia

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REGULATORY T-CELLS IN CHRONIC LYMPHOCYTIC LEUKEMIA

Mediterranean Journal of Hematology and Infectious Diseases ◽

10.4084/mjhid.2012.053 ◽

2012 ◽

Vol 4 (1) ◽

pp. e2012053 ◽

Cited By ~ 9

Author(s):

Giovanni D'arena ◽

Giovanni Rossi ◽

Barbara Vannata ◽

Silvia Deaglio

Keyword(s):

T Cells ◽

Chronic Lymphocytic Leukemia ◽

Regulatory T Cells ◽

Disease Status ◽

Lymphocytic Leukemia ◽

Small Subset ◽

Systemic Lupus Erythematosis ◽

Cell Frequency ◽

Self Tolerance

Regulatory T-cells (Tregs) constitute a small subset of cells that are actively involved in maintaining self-tolerance, in immune homeostasis and in antitumor immunity. They are thought to play a significant role in the progression of cancer and are generally increased in patient with chronic lymphocytic leukemia (CLL). Their number correlates with more aggressive disease status and is predictive of the time to treatment, as well. Moreover, it is now clear that dysregulation in Tregs cell frequency and/or function may result in a plethora of autoimmune diseases, including multiple sclerosis, type 1 diabetes mellitus, myasthenia gravis, systemic lupus erythematosis, autoimmune lymphoproliferative disorders, rheumatoid arthritis, and psoriasis. Efforts are made aiming to develop approaches to deplete Tregs or inhibit their function in either cancer and autoimmune disorders.

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