Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ligand binding of the B-cell lineage antigen CD40 enhances growth and interleukin-6 (IL-6) secretion in human B cells (the CD40/IL-6 loop). IL-6 has an autocrine and paracrine role in human multiple myeloma (MM) cell growth. With the use of the CD40 monoclonal antibody (MoAb) G28-5, we examined CD40 expression and the effect of CD40 binding on MM clonogenic colony (MCC) formation to characterize the IL-6/CD40 loop activity in MM. CD40 was expressed on plasmacytoid cells in 21 of 28 plasma cell dyscrasia (PCD) bone marrow (BM) biopsies tested (10 of 14 MM, 2 of 2 Waldenstrom's macroglobulinemia [WM], 2 of 2 plasma cell leukemia [PCL], 6 of 8 monoclonal gammopathy of undetermined significance [MGUS], and 1 of 2 primary amyloidosis [AL]). G28-5 binding increased MCCs by 35% to 150% in 11 of 17 CD40+ PCD BM cultures, but did not affect MCC formation in CD40- specimens or normal BM colony forming units (CFU-GEMM, CFU-GM, BFU-E). Responsive cultures originated from BM of patients with MM (2 of 5 cases tested), WM (2 of 2), PCL (2 of 2), and MGUS (5 of 6). CD40-responsiveness was not significantly inhibited by the presence of an anti-IL-6 MoAb (2 of 2 MGUS cultures tested), and did not correlate with the capacity to respond to IL-6 stimulation (n = 17, P > .05) or a detectable level of endogenous IL-6 (n = 15, P > .05). Additional studies were performed with PCD cell lines to characterize the interrelationship of CD40 activation and IL-6 production. Fifty percent to greater than 95% of cells from the RPMI 8226 and ARH77 lines expressed CD40, whereas 6% of U266 cells were CD40+. For RPMI 8226, ARH-77, and U266 cells, the increased MCC formation after anti-CD40 stimulation was not affected by the presence of an anti-IL-6 neutralizing MoAb and was not accompanied by detectable IL-6 secretion. There was no apparent increase in IL-6 mRNA transcription following G28-5 treatment of U266 or RPMI 8226 cells. Our observations indicate that CD40 is expressed in a subset of human myeloma cells present in various PCDs. Cell-line studies suggest that the CD40+ myeloma cell may regulate MM clonogenic colony formation without activating the IL-6 pathway.
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PMID:Anti-CD40 antibody binding modulates human multiple myeloma clonogenicity in vitro. 752 65

Spontaneous apoptosis in germinal center (GC) B cells can be arrested either by engaging cell surface immunoglobulin (Ig) with immobilized ligand or, more effectively, by treatment with soluble monoclonal antibody (mAb) directed against CD40. The present study examines the intracellular signal transduction pathways through which rescue from spontaneous apoptosis is engendered in GC B cells following ligation of surface CD40. Cross-linking the surface CD40 of GC B cells with mAb consistently resulted in enhanced tyrosine phosphorylation on a number of distinct substrates: this process could be blocked, in a dose-dependent fashion, by pre-treating GC B cells with the selective protein tyrosine kinase(s) (PTK) inhibitor, herbimycin A. Moreover, the pattern of phosphorylation on tyrosine observed following treatment with anti-CD40 was remarkably similar to that triggered by polyvalent anti-Ig. By contrast, anti-CD40 failed to stimulate the increase in inositol 1,4,5-trisphosphate and cytosolic free calcium observed in both GC B cells and resting B lymphocytes following ligation of surface Ig. The involvement of the signaling pathways generated in the rescue of GC B cells from apoptosis was studied by using selective inhibitors of PTK and of extracellular and intracellular Ca2+. Pre-incubation with the PTK inhibitor herbimycin A (5 microM) abrogated anti-CD40-mediated rescue of GC B cells from apoptosis, while genistein (40 microM) and the tyrphostins AG490 (10 microM) and AG814 (25 microM) significantly inhibited this process. Consistent with these results, herbimycin A (5 microM) abolished the expression of the 26 kDa bcl-2 protooncogene product, which confers resistance to apoptosis, normally observed following culture with anti-CD40. The Ca2+ chelators BAPTA and EGTA did not significantly affect CD40-promoted rescue. Taken together, these results indicate that CD40 of GC B cells is coupled to functional PTK but not to the phosphatidylinositol signaling pathway and that tyrosine phosphorylation is mandatory for CD40-mediated rescue of GC B cells from apoptosis.
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PMID:Protein tyrosine phosphorylation is mandatory for CD40-mediated rescue of germinal center B cells from apoptosis. 769 10

In vitro studies have indicated that chronic lymphocytic leukemia of B-cell origin (B-CLL) is resistant to cytotoxic effector lymphocytes such as natural killer and lymphokine activated killer (LAK) cells. We show here that B-cell cells are sensitive to Staphylococcal enterotoxin (SE) A-directed T-cell killing. Activation of the target cells by phorbol ester (tetradecanoyl phorbol acetate, [TPA]) greatly enhances their sensitivity to lysis. In SE-dependent cellular cytotoxicity (SDCC), members of the SE superantigen family form a bridge between T cells and target cells expressing major histocompatability complex class II molecules. Binding of SEA to the T-cell-receptor V beta region induces a strong cytotoxic capacity and cytokine production. Cells from 9 B-CLL patients were cultured in the presence or absence of TPA and used as targets in a 4-hour SDCC assay using an allogeneic T-cell line as effector. At an effector:target cell ratio 30:1, 70% to 80% of TPA-induced B-CLL cells were killed. Even at the effector:target ratio of 3:1, 47% +/- 6% of TPA-activated B-cell cells were lysed compared with 13% +/- 2% of resting cells (P < .001). A T-cell line established from a B-CLL patient killed autologous tumor cells as efficiently as allogeneic effectors. SEA-directed T cells were far more lytic to B-CLL cells compared with LAK cells or lectin (phytohemagglutinin-directed T cells. Mechanisms of SDCC lysis were investigated. Effector plus target cell supernatants contained high levels of tumor necrosis factor (TNF)-alpha and interferon-gamma, but these supernatants were not directly toxic to B-CLL cells in short term culture. High concentrations of recombinant TNF-alpha or TNF-beta had no lytic effect. Addition of neutralizing anti-TNF-alpha and anti-TNF-beta antibodies into the SDCC assay did not inhibit SEA-directed T-cell killing. TPA-activated B-CLL cells showed a 1.2- to 13-fold increased expression of the adhesion molecules intercellular adhesion molecule-1 (ICAM-1), lymphocyte function-associated antigen (LFA)-1, and LFA-3, whereas expression of HLA class II molecules increased up to 5 times. The expression of CD72, CD40, and BB-1/B7 increased 1.8 to 4.5 times. The role of these surface molecules in SDCC was analyzed in blocking experiments with monoclonal antibodies. Antibodies to ICAM-1, CD18, and HLA-DR abolished the cytotoxicity, and a substantial reduction was seen with antibody to CD72.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Efficient killing of chronic B-lymphocytic leukemia cells by superantigen-directed T cells. 810 59

The in vitro analysis of growth regulation in low-grade B non-Hodgkin's lymphoma (B-NHL) is hampered by the rapid apoptotic death of the malignant B cells ex vivo. A complex culture system, using murine CDw32 transfected fibroblasts (LTK-cells), IL-4 and anti-CD40 mAb, has been established for the propagation of normal mature B cells in vitro. We investigated the influence of the different components of this coculture system on cell survival and apoptosis of B-NHL cells. Nine samples from patients with follicular lymphoma and from eight patients with immunocytoma were analyzed. No cell proliferation of B-NHL cells could be induced in the culture system. However, CDw32-transfected murine fibroblasts most efficiently supported cell viability of B-NHL cells with an increase in cell survival by 114% compared to the control (P = 0.047). IL-4 alone also had a stimulatory effect on cell survival of B-NHL cells after 6 days. In contrast, the soluble recombinant CD40 ligand gp39 and the anti-CD40 mAbs mAb89 and EA-5 did not prolong cell survival. CDw32 transfectants blocked apoptosis of B-NHL cells efficiently from 67% in the control to 16% (P = 0.001). Reduction in apoptosis was accompanied by an elevated bcl-2 protein expression. IL-4 or mAb89 did not further reduce apoptotic cell death in CDw32 transfectant-dependent cocultures. Our data underline the pivotal role of LTK- cells for cell survival of B-NHL cells in vitro. The efficient blockage of apoptosis associated with increased bcl-2 protein expression causes prolonged cell viability of the B-NHL cells.
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PMID:In vitro activation of low-grade non-Hodgkin's lymphoma by murine fibroblasts, IL-4, anti-CD40 antibodies and the soluble CD40 ligand. 936 19

OX40 ligand (OX40L), a member of the TNF family, was shown to be capable of signaling both the cells on which it is expressed and those expressing OX40, its cognate receptor. Here we show that OX40L is expressed on dendritic cells (DC), the most efficient APC to prime naive T cells. The expression and the functional activity of OX40L were examined by means of mAbs used to stain or cross-link OX40L on 1) freshly isolated human blood DC (bDC) and 2) monocyte-derived DC at different stages of differentiation. These were derived from monocytes cultured either with IL-4 and granulocyte-macrophage CSF (IL-4-Mo-DC) or with IL-4 and granulocyte-macrophage CSF plus TNF-alpha. Both types of Mo-DC expressed OX40L after stimulation through CD40; ligation of OX40L on activated IL-4-Mo-DC enhanced by 4- to 35-fold their cytokine production (TNF-alpha, IL-12 p40, IL-1 beta, and IL-6) and increased CD80, CD86, CD54, and CD40 expression. Stimulation of activated IL-4-Mo-DC through OX40L strikingly enhanced their maturation as evidenced by CD83 up-regulation, CD115 (CSF-1R) down-regulation, and typical morphologic changes. OX40L was constitutively expressed on a subset of bDC, and its ligation slightly enhanced CD40L-stimulated IL-12 production. OX40L was down-regulated after overnight culture and spontaneously reexpressed on a subset of mature bDC (CD83high, CD33high, CD11chigh, CD5+). Thus, the expression of OX40L on DC suggests a physiologic role of this molecule during T cell priming by virtue of its ability to costimulate both T cell and DC activation and differentiation.
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PMID:Expression and function of OX40 ligand on human dendritic cells. 937 71

Peptides are flexible molecules and can adopt local structural features of protein, such as secondary structure, hydrophobicity, and distribution of electrostatic charges, and so forth, and mimic their functions. Therapeutic peptidomimetics that are immunologically relevant are developed by engineering the surface loop structures in the proteins and receptors. The class of molecules targeted include immunoglobulin fold-containing molecules: antibody, cell-surface CD4 receptors and cystine-knot-containing receptor family members: tumor necrosis factor (TNF), CD40, and p185/Neu receptors. We have used the loops involved in the molecular recognition as a template and developed peptidomimetics that interfere with the functions of the target molecules. In this article, two molecular targets are discussed: (1) immunoglobulin fold-containing CD4 receptor and (2) cystine-knot-containing TNF receptor (TNFR).
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PMID:Structure-based design of immunologically active therapeutic peptides. 947 78

Ligation of CD40 on monocytes through its interaction with CD40 ligand (CD154) present on activated T helper cells, results in activation of monocyte inflammatory cytokine synthesis and rescue of monocytes from apoptosis induced through serum deprivation. Both of these consequences of CD40 stimulation have been shown to be dependent on the induction of protein tyrosine kinase activity. CD40-mediated activation of protein tyrosine kinase activity and subsequent inflammatory cytokine production are abrogated by treatment of monocytes with the T helper type 2 cytokines interleukin 4 (IL-4) and interleukin 10 (IL-10). In the current study we demonstrate that stimulation of monocytes through CD40 resulted in the phosphorylation and activation of the extracellular signal-regulated kinases 1 and 2 (ERK1/2) mitogen-activated protein kinases, whereas phosphorylation of mitogen-activated protein kinases family members p38 and c-Jun N-terminal kinase was not observed in response to this stimuli over the time course examined. PD98059, an inhibitor of the upstream activator of ERK1/2, the MAP/ERK kinase MEK1/2, suppressed IL-1beta and tumor necrosis factor-alpha production in a dose-dependent fashion. Pretreatment of monocytes with IL-4 and IL-10 inhibited CD40-mediated activation of ERK1/2 kinase activity when used individually, and are enhanced in effectiveness when used in combination. Together, the data demonstrate that CD40-mediated induction of IL-1beta and tumor necrosis factor-alpha synthesis is dependent on a MEK/ERK pathway which is obstructed by signals generated through the action of IL-4 and IL-10.
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PMID:CD40 signaling of monocyte inflammatory cytokine synthesis through an ERK1/2-dependent pathway. A target of interleukin (il)-4 and il-10 anti-inflammatory action. 1002 6

Follicular dendritic cells (FDC)3 play crucial roles in germinal center (GC) formation and differentiation of GC B cells. Many aspects of FDC function are influenced by contact with B or T cells, and by cytokines produced in the GC, which involve stimulation of CD40 and TNF-alpha receptors on FDC. In this study, using an established FDC line, HK cells, we compared the effects of CD40 and TNF receptor triggering on cytokine induction and activation of mitogen-activated protein kinase family. We show that HK cells spontaneously produced IL-6, M-CSF, and G-CSF mRNA. Both the soluble form of CD40 ligand (sCD40L) and TNF increased the level of M-CSF and G-CSF mRNA. While TNF strongly induced IL-6 mRNA, its expression was not affected by sCD40L treatment, differing from the strong IL-6 induction in other cell types upon CD40 stimulation. In addition, sCD40L treatment resulted in activation of extracellular signal-related kinase 1 and 2 (ERK1/2) and p38 without significant increase in c-Jun N-terminal kinase (JNK) activity. Lack of JNK activation differs in that most B cells respond to CD40 stimulation by inducing JNK activity strongly, suggesting distinct characteristics of CD40 signaling in FDC. Compared with the effects of sCD40L, TNF was capable of inducing JNK activity in addition to the activation of ERK1/2 and p38. Furthermore, the proximal signaling elements activated by TNF differed from those activated by sCD40L, in that TNF did not require PMA-sensitive protein kinase C isoforms in the activation of ERK and p38, whereas sCD40L did. However, signals activated by these stimuli converged on cytokine gene expression in a synergistic manner, which may have implication in augmenting FDC function during GC reaction.
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PMID:Differential induction of cytokine genes and activation of mitogen-activated protein kinase family by soluble CD40 ligand and TNF in a human follicular dendritic cell line. 1039 51

Cytotoxic T lymphocytes (CTLs) can kill target cells by the granule/exocytosis pathway or the Fas-mediated apoptosis pathway. The sensitivity of chronic lymphocytic leukemia (CLL) B cells to CTL-mediated apoptosis before and after CD40 activation was examined. Resting or CD40-activated CLL cells were found to be equally sensitive to class I-restricted CTL-mediated killing. Despite expressing CD95, the CD40-activated CLL target cells were found to be resistant to apoptosis induced by CH11, an IgM CD95 monoclonal antibody (mAb). Consistent with this, inhibitors of caspases, which are involved in the Fas-induced apoptotic pathway (eg, N-carbobenzoxy-Val-Ala-Asp fluoromethyl ketone [z-VAD-fmk]), were unable to block destruction of CLL target cells by CTL. In addition, preincubation of the effector T cells with the anti-Fas ligand mAb NOK-2 failed to inhibit their subsequent ability to kill CLL target cells. On the other hand, CTL activity was blocked by inhibitors of the granule exocytosis pathway such as ethylene-glyco-tetra-acetic acid or concanamycin A. These results indicate that CD40 activation does not impair the sensitivity of CLL cells to Fas-independent CTL-mediated apoptosis. (Blood. 2000;95:3853-3858)
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PMID:CD40 activation does not protect chronic lymphocytic leukemia B cells from apoptosis induced by cytotoxic T lymphocytes. 1084 20

Despite CD40's role in stimulating dendritic cells (DCs) for efficient specific T-cell stimulation, its signal transduction components in DCs are still poorly documented. We show that CD40 receptors on human monocyte-derived DCs associate with sphingolipid- and cholesterol-rich plasma membrane microdomains, termed membrane rafts. Following engagement, CD40 utilizes membrane raft-associated Lyn Src family kinase, and possibly other raft-associated Src family kinases, to initiate tyrosine phosphorylation of intracellular substrates. CD40 engagement also leads to a membrane raft-restricted recruitment of tumor necrosis factor (TNF) receptor-associated factor (TRAF) 3 and, to a lesser extent, TRAF2, to CD40's cytoplasmic tail. Thus, the membrane raft structure plays an integral role in proximal events of CD40 signaling in DCs. We demonstrate that stimulation of Src family kinase within membrane rafts initiates a pathway implicating ERK activation, which leads to interleukin (IL)-1alpha/beta and IL-1Ra mRNA production and contributes to p38-dependent IL-12 mRNA production. These results provide the first evidence that membrane rafts play a critical role in initiation of CD40 signaling in DCs, and delineate the outcome of CD40-mediated pathways on cytokine production.
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PMID:CD40 signaling in human dendritic cells is initiated within membrane rafts. 1088 Apr 43


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