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PubMed1996

Platelet-derived growth factor activates protein kinase C epsilon through redundant and independent signaling pathways involving phospholipase C gamma or phosphatidylinositol 3-kinase.

Protein kinase C (PKC), a major cellular receptor for tumor-promoting phorbol esters and diacylglycerols (DGs), appears to be involved in a variety of cellular functions, although its activation mechanism in vivo is not yet fully understood. To evaluate the signaling pathways involved in the activation of PKC epsilon upon stimulation by platelet-derived growth factor (PDGF) receptor (PDGFR), we used a series of PDGFR "add-back" mutants. Activation of a PDGFR mutant (Y40/51) that binds and activates phosphatidylinositol 3-kinase (PI 3-kinase) caused translocation of PKC epsilon from the cytosol to the membrane in response to PDGF. A PDGFR mutant (Y1021) that binds and activates phospholipase C gamma (PLC gamma), but not PI 3-kinase, also caused the PDGF-dependent translocation of PKC epsilon. The translocation of PKC epsilon upon stimulation of PDGFR (Y40/51) was inhibited by wortmannin, an inhibitor of PI 3-kinase. Activation of PKC epsilon was further confirmed in terms of PKC epsilon-dependent expression of a phorbol 12-tetradecanoate 13-acetate response element (TRE)-luciferase reporter. Further, purified PKC epsilon was activated in vitro by either DG or synthetic phosphatidylinositol 3,4,5-trisphosphate. These results clearly demonstrate that PKC epsilon is activated through redundant and independent signaling pathways which most likely involve PLC gamma or PI 3-kinase in vivo and that PKC epsilon is one of the downstream mediators of PI 3-kinase whose downstream targets remain to be identified.

Cell Compartmentation↗
PubMed2001

Recombinant perciform GnRH-R activates different signaling pathways in fish and mammalian heterologous cell lines.

Perciforms have three forms of gonadotropin-releasing hormone (GnRH) in their brain. All three GnRHs are potent secretogogues for luteinizing hormone (LH) from the pituitary. The pivotal role of GnRH-R-GnRH interactions in reproductive homeostasis is well established; however, there is a paucity of information on how a GnRH-R responds to the three endogenous GnRH forms in a perciform species. In this study, a recombinant pituitary GnRH-R from striped bass (stb) was expressed in a mammalian cell line (COS-7) and a fish cell line (CHSE-214). Activation of the signaling pathways was monitored by reporter gene (luciferase) based assays, which were specific for cAMP-PKA or Ca 2+/calmodulin kinase (activated via c-fos promoter) signaling pathways. The stbGnRH-R expressed in two different cell lines triggered different downstream signaling in response to the treatments with chicken (c) GnRH II. Interestingly, when endogenous GnRHs were used in combinations, the luciferase activity was significantly attenuated in transfected CHSE-214 cells.

Animals↗
PubMed2003

Neuroglian activates Echinoid to antagonize the Drosophila EGF receptor signaling pathway.

echinoid (ed) encodes an cell-adhesion molecule (CAM) that contains immunoglobulin domains and regulates the EGFR signaling pathway during Drosophila eye development. Based on our previous genetic mosaic and epistatic analysis, we proposed that Ed, via homotypic interactions, activates a novel, as yet unknown pathway that antagonizes EGFR signaling. In this report, we demonstrate that Ed functions as a homophilic adhesion molecule and also engages in a heterophilic trans-interaction with Drosophila Neuroglian (Nrg), an L1-type CAM. Co-expression of ed and nrg in the eye exhibits a strong genetic synergy in inhibiting EGFR signaling. This synergistic effect requires the intracellular domain of Ed, but not that of Nrg. In addition, Ed and Nrg colocalize in the Drosophila eye and are efficiently co-immunoprecipitated. Together, our results suggest a model in which Nrg acts as a heterophilic ligand and activator of Ed, which in turn antagonizes EGFR signaling.

Animals↗
PubMed1999

Mitogen-activated protein kinase and protein kinase A signaling pathways stimulate cholecystokinin transcription via activation of cyclic adenosine 3',5'-monophosphate response element-binding protein.

Cholecystokinin (CCK) is a potent neuropeptide expressed in the small intestine and in the central nervous system. We have examined the effect of basic fibroblast factor (bFGF) and forskolin on CCK gene transcription and depicted the signaling pathways that lead to promoter activation. bFGF and forskolin stimulated promoter activity via a cAMP response element (CRE)/12-O-tetradecanoylphorbol-13-acetate response element (TRE) located 80 bp upstream from the transcription initiation site. In nuclear extracts from unstimulated as well as stimulated cells, only CRE-binding protein (CREB) and activating transcription factor-1 (ATF-1) bound to the CRE/TRE, and activation was associated with phosphorylation of CREB serine-133 and ATF-1 serine-63. In murine F9 cells, CREB stimulated promoter activity 10-fold in the presence of protein kinase A (PKA), and in SK-N-MC cells activation was inhibited 60-70% by a dominant negative CREB mutant. In contrast, ATF-1 had no effect in F9 cells and exhibited a dominant negative effect in SK-N-MC cells. bFGF stimulation led to phosphorylation of the p38 mitogen-activated protein kinase (MAPK), and the extracellular signal-regulated kinase (ERK) MAPK and promoter activation, phosphorylation of CREB, and GAL4-CREB-dependent transcription were selectively prevented by a dominant negative Ras-mutant, the p38 MAPK-specific inhibitor SB203580, and the MAP/ERK kinase 1 (MEK1) inhibitor PD098059. Forskolin stimulation proceeded via the PKA pathway, and to a minor extent via the p38 and ERK MAPK pathways. We conclude that bFGF and forskolin stimulate the CCK gene promoter via the CRE/TRE(-80) in the proximal promoter region. Signaling proceeds through the p38 MAPK, the ERK MAPK, and the PKA-signaling pathways, which leads to cumulative phosphorylation and activation of CREB. We propose that bFGF in combination with neurotransmitters/neuropeptides coupling to the PKA-signaling pathway play an important role in the control of CCK gene expression.

Activating Transcription Factor 1↗
PubMed2006

Involvement of ERK and CREB signaling pathways in the protective effect of PACAP in monosodium glutamate-induced retinal lesion.

Pituitary adenylate cyclase-activating polypeptide (PACAP) has well-documented neuroprotective actions, which have also been shown in retinal degeneration induced by monosodium glutamate (MSG) in neonatal rats. The aim of this article was to investigate the activation of extracellular signal-regulated kinase (ERK1/2) and cyclic adenosine 3',5'-phosphate (cAMP)-responsive element binding protein (CREB) signaling pathways by Western blot analysis in retinal degeneration induced by MSG. We found that intravitreal administration of PACAP preceding the MSG treatments induced significant increases in the phosphorylation, that is, the activation of ERK1/2 and its downstream target, CREB, 12 h after the treatment compared to the contralateral untreated eye during the first two treatments, with no further elevations 24 h after treatments. These results demonstrate that the degenerative effect of MSG and the protective effect of PACAP involve complex kinase signaling pathways and are related to cAMP/ERK/CREB activation.

Animals↗
PubMed2003

Regulation of MAPK signaling pathways through immunophilin-ligand complex.

It is well established that the immunosuppressive effects of cyclosporin A (CsA) and FK506 (also known as tacrolimus) are mediated through binding to their cognate cellular proteins cyclophilin and FKBP (collectively termed immunophilins), respectively. Biochemical analysis had revealed that cyclophilin-CsA and FKBP-FK506 complexes bind to and inactivate Ca(2+)-dependent serine/threonine phosphatase calcineurin. Since calcineurin regulates nuclear translocation and subsequent activation of nuclear factor of activated T cells (NFAT) transcription factors that is one of essential steps for cytokine gene expression in activated T cells, it is believed that inhibition of calcineurin is a molecular basis of the immunosuppressive properties of CsA and FK506. However, recent studies indicate that both CsA and FK506 can block activation of JNK and p38 signaling pathways during T cell activation. CsA and FK506, thus, have two distinct mechanisms of action; one is the inhibition of the protein phosphatase activity of calcineurin, leading to the blockade of the nuclear translocation of NFAT transcription factors, and the other is the suppression of JNK and p38 activation pathways. It is likely that the presence of two distinct targets in T cell activation makes CsA and FK506 highly potent immunosuppressive drugs. Here we discuss the action of immunophilin-ligand complexes on JNK and p38 activation pathways. We also argue the possibility of immunotherapeutic application targeting at JNK and p38 signaling pathways.

Animals↗
PubMed2005

Mutually exclusive subsets of BH3-only proteins are activated by the p53 and c-Jun N-terminal kinase/c-Jun signaling pathways during cortical neuron apoptosis induced by arsenite.

The c-Jun N-terminal protein kinase (JNK)/c-Jun and p53 pathways form distinct death-signaling modules in neurons that culminate in Bax-dependent apoptosis. To investigate whether this signaling autonomy is due to recruitment of particular BH3-only proteins, we searched for a toxic signal that would activate both pathways in the same set of neurons. We show that arsenite activates both the JNK/c-Jun and p53 pathways in cortical neurons, which together account for >95% of apoptosis, as determined by using the mixed-lineage kinase (JNK/c-Jun) pathway inhibitor CEP11004 and p53-null mice. Despite the coexistence of both pathways in at least 30% of the population, Bim mRNA and protein expression was increased only by the JNK/c-Jun signaling pathway, whereas Noxa and Puma mRNA and Puma protein expression was entirely JNK/c-Jun independent. About 50% of Puma/Noxa expression was p53 dependent, with the remaining signal being independent of both pathways and possibly facilitated by arsenite-induced reduction in P-Akt. However, functionally, Puma was predominant in mediating Bax-dependent apoptosis, as evidenced by the fact that more than 90% of apoptosis was prevented in Puma-null neurons, although Bim was still upregulated, while Bim- and Noxa-null neurons died similarly to wild-type neurons. Thus, the p53 and JNK/c-Jun pathways can activate mutually exclusive subclasses of BH3-only proteins in the same set of neurons. However, other factors besides expression may determine which BH3-only proteins mediate apoptosis.

Animals↗
PubMed2005

Activation of the sphingosine kinase-signaling pathway by high glucose mediates the proinflammatory phenotype of endothelial cells.

Vascular endothelial cells are key targets for hyperglycemic damage that facilitates vascular inflammation and the vasculopathy associated with diabetes mellitus. However, the mechanisms underlying this damage remain undefined. We now demonstrate that hyperglycemia induces activation of sphingosine kinase (SphK), which represents a novel signaling pathway that mediates endothelial damage under ambient high glucose conditions. SphK activity was significantly increased in aorta and heart of streptozotocin-induced diabetic rats. Interestingly, this increase in SphK activity was prevented by insulin treatment, which achieved euglycemia in the diabetic animals. Hyperglycemia-induced increase in SphK activity was also evident in endothelial cells that received long-term exposure to high glucose (22 mmol/L). Studies using a small interfering RNA strategy demonstrated that endogenous SphK1, but not SphK2, is the major isoenzyme that was activated by high glucose. In addition, an increase in SphK1 phosphorylation was detected in a protein kinase C- and extracellular signal-regulated kinase 1/2-dependent manner, which accounts for the high glucose-induced increases in SphK activity. Importantly, inhibition of SphK1 by either a chemical inhibitor (N',N'-dimethylsphingosine) or expression of a dominant-negative mutant of SphK1 (SphK(G82D)), or SphK1-specific small interfering RNA, strongly protected endothelial cells against high glucose-induced damage, as characterized by an attenuation in the expression of proinflammatory adhesion molecules, adhesion of leukocytes to endothelial cells, and nuclear factor kappaB activation. Thus, interventions that target the SphK-signaling pathway may have the potential to prevent vascular lesions under hyperglycemic conditions.

Animals↗
PubMed2004

Differential activation of MAPK signaling pathways by TGF-beta1 forms the molecular mechanism behind its dose-dependent bidirectional effects on hematopoiesis.

We have earlier reported that transforming growth factor-beta1 (TGF-beta1), a well-known inhibitor of hematopoiesis, stimulated colony formation from adult human bone marrow mononuclear cells (BM MNC) when used at low concentrations. We examined the possible molecular mechanism behind this bidirectional effect using CD34+ cells isolated from human BM for clonal assays and the KG1a cell line as a model system for analysis of proteins for signaling pathways by immunoblotting. We found that TGF-beta1 at low doses (picogram levels) stimulated the colony formation from CD34+ cells, indicating that these progenitors form the direct target of stimulatory action of TGF-beta1. CD34+ cells were found to be more sensitive to the TGF-beta1 concentration than the total MNC. We used the KG1a cell line as a model system for identification of mitogen-activated protein kinase (MAPK) and AKT signaling pathways involved in the process. Low doses strongly induced p44/42 MAPK phosphorylation, whereas high doses induced p38 activation. Use of specific p44/42 MAPK inhibitor PD 98059 in the colony assay abrogated the stimulatory effect of low TGF-beta1. On the other hand, use of p38 MAPK inhibitor SB 203580 along with low TGF-beta1 concentrations had a synergistic effect on stimulation of colony formation. Treatment of BM MNC with Anisomycin, which activates stress kinases, resulted in a dose-dependent inhibition of colony formation. This inhibition could not be rescued by stimulatory doses of TGF-beta1. Phosphorylation of AKT was found to occur in a dose-dependent way but declined slightly at the highest concentration used (10 ng/ml). Inhibition of the AKT pathway by LY 294002 strongly suppressed colony formation. These data indicate clearly that sustained activation of p44/42 MAPK perhaps forms the stimulatory signal induced by low TGF-beta1, whereas activation of p38 forms the inhibitory pathway.

Anisomycin↗
PubMed2005

Genistein-induced apoptosis via Akt signaling pathway in anaplastic large-cell lymphoma.

More than half of anaplastic large-cell lymphoma (ALCL) are associated with chromosomal translocation t(2;5)(p23;q35) that leads to the expression of nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) oncoprotein. NPM-ALK activates the antiapoptotic phosphatidylinositol-3 kinase/Akt (PI3K/Akt) signaling pathway, which plays a critical role in cell survival and apoptosis. Inhibition of the PI3K/Akt pathway has been considered as a therapeutic target for cancer where PI3K/Akt activation is a causative factor. Genistein, a natural isoflavonoid found in soy products, has been shown to inhibit cell growth and induce apoptosis in a wide variety of cell lines. Here, we demonstrated that treatment of two t(2;5) ALCL cell lines, SUDHL-1 and Karpas299, with genistein induced apoptosis in a time- and dose-dependent manner. Concurrently, these cells exhibited a decrease in Akt protein levels and subsequent downregulation of Akt activity (Akt phosphorylation). Furthermore, genistein treatment induced mitochondrial membrane potential change, caspase-3 activation and PARP cleavage. From these results, we conclude that inhibition of the Akt signaling pathway and induction of apoptosis by genistein could be used as a new treatment modality for the prevention and/or treatment of t(2;5) ALCL and other hematopoietic malignancies.

Antineoplastic Agents↗
PubMed2002

XK469, a novel antitumor agent, inhibits signaling by the MEK/MAPK signaling pathway.

PURPOSE: XK469 (NSC 697887) is a novel antitumor agent with broad activity against a variety of tumors including drug-resistant tumors. Previous studies have indicated that XK469 is an antiproliferative agent with a low cytotoxic effect in human H116 tumor cells. In this study, we sought to determine the signaling pathways involved in mediating its antiproliferative activity. METHODS: The antiproliferative activity of XK469 was tested using human U-937 leukemia cells in culture. XK469-induced cell cycle arrest was determined using flow cytometric analysis. Phosphorylation/activation of MEK and MAPK was analyzed using immunoblot analyses with specific antibodies against p-MEK and p-MAPK. RESULTS: Cell cycle analysis revealed that XK469 arrested U-937 cells at the G(2)/M phase. Compared with conventional anticancer agents, XK469 showed very low, if any, cytotoxic or proapoptotic effect against U-937 cells. In contrast, treatment of U-937 cells with vinblastine, doxorubicin and m-AMSA resulted in extensive cell death through apoptotic pathways. XK469, but not other agents, potently inhibited the phosphorylation/activation of MEK in U-937 cells cultured in serum-containing medium. XK469 was also able to block the activation of MEK by serum addition in starved U-937 cells. Exposure of cells to XK469 for 1 h was sufficient to inhibit the activation of MEK and its downstream kinase, MAPK. The antiproliferative response to XK469 was correlated with a steady accumulation of cyclins B1 and A, which appeared to be a direct result of G(2)/M arrest. CONCLUSIONS: Our findings suggest that the antiproliferative effect of XK469 is mediated by inhibiting the MEK/MAPK signaling pathways in U-937 human leukemia cells.

Antineoplastic Agents↗
PubMed1999

H2O2 is required for UVB-induced EGF receptor and downstream signaling pathway activation.

Ultraviolet radiation (UVR)-induced receptor phosphorylation is increasingly recognized as a widely occurring phenomenon. However, the mechanisms, mediators, and sequence of events involved in this process remain ill-defined. We have recently shown that exposure of human keratinocytes to physiologic doses of ultraviolet B radiation (UVB) activates epidermal growth factor receptor (EGFR)/extracellular-regulated kinase 1 and 2 (ERK1/2), and p38 signaling pathways via reactive oxygen species. Here we demonstrate that UVB exposure increased intra- and extracellular H2O2 production rapidly in a time-dependent manner. An EGFR-specific monoclonal antibody abrogated EGFR autophosphorylation and markedly decreased the phosphorylation of ERK1/2 whereas p38 activation was unaffected. Overexpression of catalase strongly inhibited UVB-induced EGFR/ERK1/2 pathway activation. These findings establish the sequence of events after UVB irradiation: (i) H2O2 generation, (ii) EGFR phosphorylation, and (iii) ERK activation. Our results identify UVB-induced H2O2 as a second messenger that is required for EGFR and dependent downstream signaling pathways activation.

Catalase↗
PubMed2006

Cell-based assay strategy for identification of motif-specific RANK signaling pathway inhibitors.

Osteoclasts, the principal bone-resorbing cells, not only play a pivotal role in skeletal development and maintenance but are also implicated in the pathogenesis of various bone disorders such as postmenopausal osteoporosis, bone erosion in inflammatory conditions, and tumor-induced osteolysis. As a result, several antiresorptive drugs (agents capable of inhibiting osteoclast formation and/or function) have been developed and are widely used to prevent and treat these bone diseases. However, current antiresorptive agents either lack satisfactory efficacy or cause serious side effects in clinical management of these bone disorders. Almost a decade ago, the receptor activator of nuclear factor-kappaB (RANK) ligand (RANKL) was identified as an essential factor required for osteoclast formation. RANKL exerts the effect by binding to its receptor RANK on osteoclast precursors. RANKL also has a decoy receptor, osteoprotegerin (OPG), which inhibits RANKL function by competing with RANK for RANKL. The unraveling of the critical role for the RANKL/RANK/OPG system in osteoclast biology provides an unprecedented opportunity to develop more effective antiresorptive drugs. Unfortunately, the agents currently under development, such as OPG, RANK-Fc, and anti-RANKL antibodies, all inherit a serious drawback--lack of specificity, due to the involvement of the RANKL/RANK/OPG system in other biological processes such as immune response and mammary gland development. Thus, future efforts may need to shift to explore RANK signaling pathways as more effective therapeutic targets. Here, we review our current understanding of RANK signaling in osteoclasts and then discuss the potential of RANK signaling pathways as therapeutic pathways. Moreover, we further describe a strategy for constructing novel cell-based systems for identifying compounds inhibiting signaling from two recently identified RANK motifs through high throughput screening. We hope that this review will not only provide readers with an update on progress in this area of research but, more importantly, will also serve as a starting point for further discussion and eventual development of new strategies for harnessing the ultimate potential of the RANKL/RANK/OPG system as antiresorptive therapeutic targets.

Amino Acid Sequence↗
PubMed2005

Characterization of the signaling pathway downstream p75 neurotrophin receptor involved in beta-amyloid peptide-dependent cell death.

The accumulation of beta-amyloid (Abeta) peptide is a key pathogenic event in Alzheimer's disease. Previous studies have shown that Abeta peptide can damage neurons by activating the p75 neurotrophin receptor (p75NTR). However, the signaling pathway leading to neuronal cell death is not completely understood. By using a neuroblastoma cell line devoid of neurotrophin receptors and engineered to express either a full-length or a death domain (DD)-truncated form of p75NTR, we demonstrated that Abeta peptide activates the mitogen-activated protein kinases (MAPKs) p38 and c-Jun N-terminal kinase (JNK). We also found that Abeta peptide induces the translocation of nuclear factor-kappaB (NF-kappaB). These events depend on the DD of p75NTR. Beta-amyloid (Abeta) peptide was found not to be toxic when the above interactors were inhibited, indicating that they are required for Abeta-induced neuronal cell death. p75 neurotrophin receptor (p75NTR)-expressing cells became resistant to Abeta toxicity when transfected with dominant-negative mutants of MAPK kinases 3, 4, or 6 (MKK3, MKK4, or MKK6), the inhibitor of kappaBalpha, or when treated with chemical inhibitors of p38 and JNK. Furthermore, p75NTR-expressing cells became resistant to Abeta peptide upon transfection with a dominant-negative mutant of p53. These results were obtained in the presence of normal p38 and JNK activation, indicating that p53 acts downstream of p38 and JNK. Finally, we demonstrated that NF-kappaB activation is dependent on p38 and JNK activation. Therefore, our data suggest a signaling pathway in which Abeta peptide binds to p75NTR and activates p38 and JNK in a DD-dependent manner, followed by NF-kappaB translocation and p53 activation.

Amyloid beta-Peptides↗
PubMed2006

Regulation of cardiac growth and coronary angiogenesis by the Akt/PKB signaling pathway.

Postnatal growth of the heart is primarily achieved through hypertrophy of individual myocytes. Cardiac growth observed in athletes represents adaptive or physiological hypertrophy, whereas cardiac growth observed in patients with hypertension or valvular heart diseases is called maladaptive or pathological hypertrophy. These two types of hypertrophy are morphologically, functionally, and molecularly distinct from each other. The serine/threonine protein kinase Akt is activated by various extracellular stimuli in a phosphatidylinositol-3 kinase-dependent manner and regulates multiple aspects of cellular functions including survival, growth and metabolism. In this review we will discuss the role of the Akt signaling pathway in the heart, focusing on the regulation of cardiac growth, contractile function, and coronary angiogenesis. How this signaling pathway contributes to the development of physiological/pathological hypertrophy and heart failure will also be discussed.

Adaptation, Physiological↗
PubMed2004

Thyrotropin releasing hormone inhibits tau phosphorylation by dual signaling pathways in hippocampal neurons.

Depletion of thyrotropin releasing hormone (TRH) gene expression resulted in augmented tau and glycosynthetase kinase-3beta (GSK-3beta), in contrast, TRH administration resulted in decreases of 75% in GSK-3beta and 90% in Tau phosphorylation in cultured rat hippocampal neurons. To further study TRH regulation of tau phosphorylation, immunoblotting was used to explore G-protein coupled TRH receptor activation of the phosphokinase C (PKC) and phosphokinase A (PKA) signaling pathways. TRH was found to rapidly activate PKA (2.5 fold in 10 min) while it suppressed PKC (levels decreased by 85% vs. control) in hippocampal neurons. This process was also discovered to be a cell type-specific response, as TRH activated PKC in only hypothalamic neurons. Further investigation revealed that the Src inhibitor Protein Phosphatase 2 (PP2, 50 uM) could block TRH inhibition of PKC, GSK-3beta, and tau phosphorylation with no effects on PKA. In addition, the PKC inhibitor GF109203 Bis (10 uM) was also able to suppress TRH inhibition of GSK-3beta, leading to increased GSK-3 beta activity. Independent of these effects, inhibition of PKA by H89 (10 uM) significantly blocked TRH inhibition of GSK-3 beta. These data suggests that both PKA and PKC are independently crucial to TRH's effects on GSK-3 beta, and support the roles of two distinct pathways involving suppression of PKC via the Src kinase and activation of PKA in mediating TRH effects on GSK-3 beta and tau. These dual signaling pathways between TRH and tau may provide mechanisms for the precise regulation of tau phosphorylation and dephosphorylation in neurons.

Cyclic AMP-Dependent Protein Kinases↗
PubMed2001

Tyrosine kinase-dependent, phosphatidylinositol 3'-kinase, and mitogen-activated protein kinase-independent signaling pathways prevent lung adenocarcinoma cells from anoikis.

Normal epithelial cells are anchorage-dependent. Detachment of normal epithelial cells from their substratum causes apoptosis, termed anoikis. Malignant tumor cells, however, can survive and proliferate independent of anchorage. To understand the molecular basis of tumor cell anchorage independence, we investigated the role of tyrosine kinases and their downstream signaling pathways in anoikis resistance of human lung adenocarcinoma cells. Four of the five lung adenocarcinoma cell lines analyzed are resistant to anoikis. Tyrosine kinase inhibitor genistein rendered three of them sensitive to anoikis. Although cell detachment induced rapid protein tyrosine dephosphorylation in Madin-Darby canine kidney cells, a nontransformed epithelial cell line, tyrosine phosphorylation of several proteins in the tumor cells is anchorage-independent. Similarly, phosphorylation of Akt and mitogen-activated protein kinase, two signaling proteins downstream of tyrosine kinases, was decreased in Madin-Darby canine kidney cells but increased in some of the tumor cells upon cell detachment. Inhibition of phosphorylation of the two proteins, however, did not induce anoikis in the tumor cells. Specific inhibitors to several known tyrosine kinases also failed to induce anoikis in these cells. These data suggest the existence of tyrosine kinase-dependent phosphatidylinositol 3'-kinase and mitogen-activated protein kinase-independent signaling pathways that function to regulate cell survival and death. Alteration in these pathways may count for the anchorage-independent survival of the lung adenocarcinoma cells and other malignant tumor cells.

Adenocarcinoma↗
PubMed2003

MEK/ERK pathway is aberrantly active in Hodgkin disease: a signaling pathway shared by CD30, CD40, and RANK that regulates cell proliferation and survival.

The mitogen-activated protein kinase (MAPK) (also called extracellular signal-regulated kinase [ERK]) pathway has been implicated in malignant transformation and in the regulation of cellular growth and proliferation of several tumor types, but its expression and function in Hodgkin disease (HD) are unknown. We report here that the active phosphorylated form of MAPK/ERK is aberrantly expressed in cultured and primary HD cells. Inhibition of the upstream MAPK kinase (also called MEK) by the small molecule UO126 inhibited the phosphorylation of ERK and demonstrated a dose- and time-dependent antiproliferative activity in HD cell lines. UO126 modulated the levels of several intracellular proteins including B-cell lymphoma protein 2 (Bcl-2), myeloid cell leukemia-1 (Mcl-1) and caspase 8 homolog FLICE-inhibitory protein (cFLIP), and induced G2M cell-cycle arrest or apoptosis. Furthermore, UO126 potentiated the activity of apoliprotein 2/tumor necrosis factor-related apoptosis-inducing ligand (APO2L/TRAIL) and chemotherapy-induced cell death. Activation of CD30, CD40, and receptor activator of nuclear kappabeta (RANK) receptors in HD cells by their respective ligands increased ERK phosphorylation above the basal level and promoted HD cell survival. UO126 inhibited basal and ligand-induced ERK phosphorylation, and inhibited ligand-induced cell survival of HD cell lines. These findings provide a proof-of-principle that inhibition of the MEK/ERK pathway may have therapeutic value in HD.

Antineoplastic Agents↗