The Immunosuppressive Agent 15-Deoxyspergualin Functions by Inhibiting Cell Cycle Progression and Cytokine Production Following Naive T Cell Activation

Abstract Cyclic adenosine monophosphate (cAMP) is a negative regulator of T-cell activation. However, the effects of cAMP on signaling pathways that regulate cytokine production and cell cycle progression remain unclear. Here, using primary human T lymphocytes in which endogenous cAMP was increased by the use of forskolin and 3-isobutyl-1-methylxanthine (IBMX), we show that increase of cAMP resulted in inhibition of T-cell receptor (TCR)/CD3 plus CD28–mediated T-cell activation and cytokine production and blockade of cell cycle progression at the G1 phase. Increase of cAMP inhibited Ras activation and phosphorylation of mitogen-induced extracellular kinase (MEK) downstream targets extracellular signal–related kinase 1/2 (ERK1/2) and phosphatidylinositol-3-kinase (PI3K) downstream target protein kinase B (PKB; c-Akt). These functional and biochemical events were secondary to the impaired activation of ZAP-70 and phosphorylation of LAT and did not occur when cells were stimulated with phorbol ester, which bypasses the TCR proximal signaling events and activates Ras. Increase of cAMP also inhibited activation of Rap1 mediated by TCR/CD3 plus CD28. Importantly, inhibition of Rap1 activation by cAMP was also observed when cells were stimulated with phorbol ester, although under these conditions Ras was activated and cells progressed into the cell cycle. Thus, TCR plus CD28–mediated activation of ERK1/2 and PKB, cytokine production, and cell cycle progression, all of which are inhibited by cAMP, require activation of Ras but not Rap1. These results indicate that signals that regulate cAMP levels after encounter of T cells by antigen will likely determine the functional fate toward clonal expansion or repression of primary T-cell responses.

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CTLA-4 ligation blocks CD28-dependent T cell activation.

Journal of Experimental Medicine ◽

10.1084/jem.183.6.2541 ◽

1996 ◽

Vol 183 (6) ◽

pp. 2541-2550 ◽

Cited By ~ 530

Author(s):

T L Walunas ◽

C Y Bakker ◽

J A Bluestone

Keyword(s):

Cell Cycle ◽

T Cell ◽

Cell Survival ◽

T Cell Activation ◽

Cell Cycle Progression ◽

Cell Activation ◽

Cell Function ◽

Receptor Expression ◽

Negative Regulator ◽

Cycle Progression

CTLA-4 is a CD28 homologue believed to be a negative regulator of T cell function. However, the mechanism of this downregulatory activity is not well understood. The present study was designed to examine the effect of CTLA-4 ligation on cytokine production, cell survival, and cell cycle progression. The results demonstrate that the primary effect of CTLA-4 ligation is not the induction of apoptosis. Instead, CTLA-4 signaling blocks IL-2 production, IL-2 receptor expression, and cell cycle progression of activated T cells. Moreover, the effect of CTLA-4 signaling was manifested after initial T cell activation. Inhibition of IL-2 receptor expression and cell cycle progression was more pronounced at late (72 h) time points after initial activation. The effects of anti-CTLA-4 mAbs were most apparent in the presence of optimal CD28-mediated costimulation consistent with the finding that CTLA-4 upregulation was CD28-dependent. Finally, the addition of exogenous IL-2 to the cultures restored IL-2 receptor expression and T cell proliferation. These results suggest that CTLA-4 signaling does not regulate cell survival or responsiveness to IL-2, but does inhibit CD28-dependent IL-2 production.

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Imatinib Mesylate Disrupts Cell Cycle Progression, Modifies the Nucleoskeleton and Suppresses Activation-Induced Transcription in Human T Cells.

Blood ◽

10.1182/blood.v104.11.2914.2914 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2914-2914

Author(s):

Allan Dietz ◽

William B. Johnson ◽

Gaylord J. Knutson ◽

Peggy A. Bulur ◽

Bertie Schulenberg ◽

...

Keyword(s):

Cell Cycle ◽

T Cells ◽

T Cell ◽

Imatinib Mesylate ◽

T Cell Activation ◽

Cell Cycle Progression ◽

Cell Activation ◽

Structural Changes ◽

Activated T Cells ◽

Cycle Progression

Abstract Imatinib mesylate (imatinib, Gleevec®, Novartis, Basel, Switzerland) inhibits T cells in vitro and in vivo (Dietz et al., Blood104: 1094–1099, 2004; Cwynarski et al., Leukemia18: 1332–1339, 2004). The drug blocks T cell cycle progression rather uniquely as it neither inhibits expression of CD69, an early marker of T cell activation, nor induces apoptosis. To characterize the molecular effects of imatinib leading to this mode of T-cell inhibition, we measured the changes in transcriptome (by Affymetrix U133 chips), proteome and phosphoproteome (by Western blotting, differential phosphoprotein expression and mass spectrometry). We found that phytohemagglutinin activated T cells pre-treated with imatinib had reduced expression of 983 transcripts and increased expression of 271 transcripts when compared to untreated PHA activated T cells by the factor of 1.5 or more (p<0.05). Among the prominently down-regulated transcripts were granzyme B, CTLA-4 and IL-2-receptor α-chain (CD25), all characteristic of activated T cells, as well as cyclins D2 and D3 and cyclin-dependent kinases 3, 4 and 7, the molecules regulating cell cycle progression. Among the up-regulated transcripts were Kruppel-like transcription factors 2 and 7, and p27, a finding compatible with the observed cell cycle inhibition. Furthermore, we selected and identified 30 proteins from 2-D gels that were up-regulated and/or hyperphosphorylated in imatinib treated activated T cells. Among these were four heterogeneous ribonucleoproteins, three lamins and γ-actin, all components of the nucleoskeleton at the interface of chromatin and inner nuclear membrane and involved in replication and transcription (Herrmann and Foisner, Cell. Mol. Life Sci.60: 1607–1612, 2003; Shumaker et al. Curr. Opinion Cell Biol.15: 358–366, 2003). Thus, imatinib-borne interference with T cell signal transduction affects the nuclear structure indicating for the first time that nucleoskeleton structural changes are associated with T cell activation status.

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Role of astrocytes in antigen presentation and naive T-cell activation

Journal of Neuroimmunology ◽

10.1016/s0165-5728(99)00215-5 ◽

2000 ◽

Vol 106 (1-2) ◽

pp. 69-77 ◽

Cited By ~ 42

Author(s):

Anne Cornet ◽

Estelle Bettelli ◽

Mohamed Oukka ◽

Christophe Cambouris ◽

Virginia Avellana-Adalid ◽

...

Keyword(s):

T Cell ◽

Antigen Presentation ◽

T Cell Activation ◽

Cell Activation ◽

Naïve T Cell ◽

Naive T Cell

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The tetraspanin CD9 is preferentially expressed on the human CD4+CD45RA+ naive T cell population and is involved in T cell activation

Clinical & Experimental Immunology ◽

10.1111/j.1365-2249.2004.02494.x ◽

2004 ◽

Vol 137 (1) ◽

pp. 101-108 ◽

Cited By ~ 23

Author(s):

H. KOBAYASHI ◽

O. HOSONO ◽

S. IWATA ◽

H. KAWASAKI ◽

M. KUWANA ◽

...

Keyword(s):

T Cell ◽

Cell Population ◽

T Cell Activation ◽

Cell Activation ◽

Naïve T Cell ◽

Naive T Cell

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Role of astrocytes in antigen presentation and naive T cell activation

Journal of Neuroimmunology ◽

10.1016/s0165-5728(98)91391-1 ◽

1998 ◽

Vol 90 (1) ◽

pp. 36

Author(s):

A. Cornet ◽

E. Betelli ◽

M. Oukka ◽

C. Cambouris ◽

K. Kosmatopoulos ◽

...

Keyword(s):

T Cell ◽

Antigen Presentation ◽

T Cell Activation ◽

Cell Activation ◽

Naïve T Cell ◽

Naive T Cell

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Generating Chimeric Antigen Receptors Utilizing Novel Anti-CD3 Nanobeads

Blood ◽

10.1182/blood.v124.21.5949.5949 ◽

2014 ◽

Vol 124 (21) ◽

pp. 5949-5949

Author(s):

Liora M. Schultz ◽

Debra K Czerwinski ◽

Aihua Fu ◽

Shoshana Levy ◽

Ronald Levy

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Activation ◽

Cell Activation ◽

Antigen Receptors ◽

Chimeric Antigen Receptors ◽

Car T Cell ◽

Car T ◽

Naïve T Cell ◽

Naive T Cell

Abstract The processes of ex vivo transduction of T cells to express chimeric antigen receptors (CARs) and of CAR+ T cell expansion influence the phenotype, function and ultimate fate of the final CAR+T cell product infused into patients. CAR constructs, despite expression of endogenous activation signals, require exogenous T cell activation during CAR transduction to allow optimal lenti-viral or retroviral-mediated integration of the CAR gene of interest into T cells. Clinical CAR therapy trials utilize anti-CD3 antibody-mediated activation or combined CD3 and CD28 stimulation using CD3, CD28 specific magnetic beads. We introduce novel magnetic nanoparticle beads generated from iron oxide nanoparticles conjugated to streptavidin and bound to biotinylated T cell activating antibodies for the purpose of CAR transduction. The small size of these nanobeads confers the advantage of decreased steric hindrance and enhanced capability of bead surface antibodies to access T cell surface antigen for binding and stimulation. We achieve efficient CAR transduction using anti-CD3 nanobead-mediated T cell stimulation and demonstrate CD19 specific CAR-mediated cytotoxicity of CD19+ tumor using an annexin V and 7AAD cytotoxicity assay. Evaluation of T cell phenotype following anti-CD3 nanobead-mediated T cell activation demonstrates preferential activation of naïve T cells as compared to central and effector memory cells. Addition of anti-CD28 costimulation is not necessary to achieving or inhibiting this preferential naïve T cell activation. Naïve T cells exhibit greater replicative capacity and anti-tumor function as compared to both effector and central memory T cells for adoptive transfer. We anticipate that preferential generation of naïve T cell derived CAR+ T cells achieved by introducing anti-CD3 nanobead stimulation can further improve the outcomes of clinical trials using CAR therapy. Disclosures Fu: NVIGEN Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.

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CTLA-4 engagement inhibits IL-2 accumulation and cell cycle progression upon activation of resting T cells.

Journal of Experimental Medicine ◽

10.1084/jem.183.6.2533 ◽

1996 ◽

Vol 183 (6) ◽

pp. 2533-2540 ◽

Cited By ~ 604

Author(s):

M F Krummel ◽

J P Allison

Keyword(s):

Cell Cycle ◽

T Cells ◽

Cell Death ◽

T Cell ◽

T Cell Activation ◽

Cell Cycle Progression ◽

Cell Activation ◽

Apoptotic Cell ◽

Activated T Cells ◽

Activated State

While interactions between CD28 and members of the B7 family costimulate and enhance T cell responses, recent evidence indicates that the CD28 homologue CTLA-4 plays a downregulatory role. The mechanism by which this occurs is not clear, but it has been suggested that CTLA-4 terminates ongoing responses of activated T cells, perhaps by induction of apoptosis. Here we demonstrate that CTLA-4 engagement by antibody cross-linking or binding to B7 inhibits proliferation and accumulation of the primary T cell growth factor, IL-2, by cells stimulated with anti-CD3 and anti-CD28. This inhibition is not a result of enhanced cell death. Rather it appears to result from restriction of transition from the G1 to the S phase of the cell cycle. Our observation that upregulation of both the IL-2R alpha chain and the CD69 activation antigen are inhibited by CTLA-4 engagement supplies further evidence that CTLA-4 restricts the progression of T cells to an activated state. Together this data demonstrates that CTLA-4 can regulate T cell activation in the absence of induction of apoptotic cell death.

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