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:2.7.11.2 (PDK1)
2,238 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chronic lymphocytic leukemia (CLL) is one of the most commonly diagnosed leukemias managed by practicing hematologists. For many years patients with CLL have been viewed as similar, with a long natural history and only marginally effective therapies that rarely yielded complete responses. Recently, several important observations related to the biologic significance of V(H) mutational status and associated ZAP-70 overexpression, disrupted p53 function, and chromosomal aberrations have led to the ability to identify patients at high risk for early disease progression and inferior survival. Concurrent with these investigations, several treatments including the nucleoside analogues, monoclonal antibodies rituximab and alemtuzumab have been introduced. Combination of these therapies in clinical trials has led to high complete and overall response rates when applied as initial therapy for symptomatic CLL. Thus, the complexity of initial risk stratification of CLL and treatment has increased significantly. Furthermore, when these initial therapies do not work, approach of the CLL patient with fludarabine-refractory disease can be quite challenging. This session will describe the natural history of a CLL patient with emphasis on important decision junctures at different time points in the disease. In Section I, Dr. Stephan Stilgenbauer focuses on the discussion that occurs with CLL patients at their initial evaluation. This includes a review of the diagnostic criteria for CLL and prognostic factors utilized to predict the natural history of the disease. The later discussion of risk stratification focuses on molecular and genomic aberrations that predict rapid progression, poor response to therapy, and inferior survival. Ongoing and future efforts examining early intervention strategies in high risk CLL are reviewed. In Section II, Drs. Ian Flinn and Jesus G. Berdeja focus on the discussion of CLL patients when symptomatic disease has developed. This includes an updated review of monotherapy trials with nucleoside analogs and recent trials that have combined these with monoclonal antibodies and/or alternative chemotherapy agents. Appropriate application of more aggressive therapies such as autologous and allogeneic immunotherapy and less aggressive treatments for appropriate CLL patient candidates are discussed. In Section III, Dr. John Byrd focuses on the discussion that occurs with CLL patients whose disease is refractory to fludarabine. The application of genetic risk stratification in choosing therapy for this subset of patients is reviewed. Available data with conventional combination based therapies and monoclonal antibodies are discussed. Finally, alternative promising investigational therapies including new antibodies, kinase inhibitors (CDK, PDK1/AKT, PKC) and alternative targeted therapies (DNA methyltransferase inhibitors, histone deacetylase inhibitors, etc.) are reviewed with an emphasis on the most promising agents for this patient population.
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PMID:Chronic lymphocytic leukemia. 1556 82

The RAS-activated RAF-->MEK-->extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3'-kinase (PI3'-kinase)-->PDK1-->AKT signaling pathways are believed to cooperate to promote the proliferation of normal cells and the aberrant proliferation of cancer cells. To explore the mechanisms that underlie such cooperation, we have derived cells harboring conditionally active, steroid hormone-regulated forms of RAF and AKT. These cells permit the assessment of the biological and biochemical effects of activation of these protein kinases either alone or in combination with one another. Under conditions where activation of neither RAF nor AKT alone promoted S-phase progression, coactivation of both kinases elicited a robust proliferative response. Moreover, under conditions where high-level activation of RAF induced G(1) cell cycle arrest, activation of AKT bypassed the arrest and promoted S-phase progression. At the level of the cell cycle machinery, RAF and AKT cooperated to induce cyclin D1 and repress p27(Kip1) expression. Repression of p27(Kip1) was accompanied by a dramatic reduction in KIP1 mRNA and was observed in primary mouse embryo fibroblasts derived from mice either lacking SKP2 or expressing a T187A mutated form of p27(Kip1). Consistent with these observations, pharmacological inhibition of MEK or PI3'-kinase inhibited the effects of activated RAS on the expression of p27(Kip1) in NIH 3T3 fibroblasts and in a panel of bona fide human pancreatic cancer cell lines. Furthermore, we demonstrated that AKT activation led to sustained activation of cyclin/cdk2 complexes that occurred concomitantly with the removal of RAF-induced p21(Cip1) from cyclin E/cdk2 complexes. Cumulatively, these data strongly suggest that the RAF-->MEK-->ERK and PI3'K-->PDK-->AKT signaling pathways can cooperate to promote G(0)-->G(1)-->S-phase cell cycle progression in both normal and cancer cells.
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PMID:Cooperative regulation of the cell division cycle by the protein kinases RAF and AKT. 1557 89

The novel synthetic retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) induces growth arrest and apoptosis in various tumor cell lines including non-small cell lung cancer (NSCLC) cells. CD437 binds retinoic acid receptor gamma (RARgamma) selectively, and can enhance receptor-dependent transcriptional activation of various genes. However, some of the effects of this retinoid on cell growth inhibition and apoptosis appear to be receptor-independent. To gain a better understanding of the mechanism by which CD437 exerts its effects, we employed a high throughput western blotting method (PowerBlottrade mark) using 760 monoclonal antibodies to compare the levels of their target cellular signaling proteins in untreated and CD437-treated NSCLC H460 cells. CD437 (1 microM, 24 h) increased the levels of 70 proteins and decreased the levels of 28 proteins. These proteins play a role in fundamental cellular processes including: DNA synthesis and repair, transcription and DNA-binding, cell cycle, apoptosis, cytoskeleton assembly, cell adhesion, endocytosis, growth and signal transduction. Some proteins identified by this approach have been implicated previously in the effect of CD437 (e.g., p53, Bax, cyclin B, CDK2). Additionally we identified proteins that are novel candidates for mediating the cellular responses to CD437 (e.g., FAF1, Bid, caspase 8, cdk1, KAP, NDR, RBBP, 53BP2, Grb2, PLCgamma1, p70s6k, PP2Cdelta, PKBalpha/AKT, PDK1, and several DNA repair enzymes).
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PMID:Identification of protein modulation by the synthetic retinoid CD437 in lung carcinoma cells using high throughput immunoblotting. 1564 34

The abilities of mutated active RAS proteins to modulate cell survival following exposure to ionizing radiation and small molecule kinase inhibitors were examined. Homologous recombination in HCT116 cells to delete the single allele of K-RAS D13 resulted in a cell line that exhibited an approximately 75% reduction in basal extracellular signal-regulated kinase 1/2, AKT, and c-jun-NH2-kinase 1/2 activity. Transfection of cells lacking K-RAS D13 with H-RAS V12 restored extracellular signal-regulated kinase 1/2 and AKT activity to basal levels but did not restore c-jun-NH2-kinase 1/2 phosphorylation. In cells expressing H-RAS V12, radiation caused prolonged intense activation of AKT. Inhibition of H-RAS V12 function, blockade of phosphatidylinositol 3-kinase (PI3K) function using small interfering RNA/small-molecule inhibitors, or expression of dominant-negative AKT abolished radiation-induced AKT activation, and radiosensitized these cells. Inhibition of PI3K function did not significantly radiosensitize parental HCT116 cells. Inhibitors of the AKT PH domain including perifosine, SH-(5, 23-25) and ml-(14-16) reduced the plating efficiency of H-RAS V12 cells in a dose-dependent fashion. Inhibition of AKT function using perifosine enhanced radiosensitivity in H-RAS V12 cells, whereas the SH and ml series of AKT PH domain inhibitors failed to promote radiation toxicity. In HCT116 H-RAS V12 cells, PI3K, PDK-1, and AKT were membrane associated, whereas in parental cells expressing K-RAS D13, only PDK-1 was membrane bound. In H-RAS V12 cells, membrane associated PDK-1 was phosphorylated at Y373/376, which was abolished by the Src family kinase inhibitor PP2. Inhibition of PDK-1 function using the PH domain inhibitor OSU-03012 or using PP2 reduced the plating efficiency of H-RAS V12 cells and profoundly increased radiosensitivity. OSU-03012 and PP2 did not radiosensitize and had modest inhibitory effects on plating efficiency in parental cells. A small interfering RNA generated against PDK1 also radiosensitized HCT116 cells expressing H-RAS V12. Collectively, our data argue that molecular inhibition of AKT and PDK-1 signaling enhances the radiosensitivity of HCT116 cells expressing H-RAS V12 but not K-RAS D13. Small-molecule inhibitory agents that blocked stimulated and/or basal PDK-1 and AKT function profoundly reduced HCT116 cell survival but had variable effects at enhancing tumor cell radiosensitivity.
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PMID:Activated forms of H-RAS and K-RAS differentially regulate membrane association of PI3K, PDK-1, and AKT and the effect of therapeutic kinase inhibitors on cell survival. 1571 97

We showed previously [K. Moissoglu, I.H. Gelman, J. Biol. Chem. 278 (2003) 47946-47959] that oncogenic v-Src could induce 7- to 10-fold greater anchorage-independent growth (AIG) in FAK-null mouse embryo fibroblasts (MEF) compared to those expressing FAK. Here, we demonstrate that the enhanced AIG (eAIG) correlates with increased activation levels of phosphatidylinositol 3-kinase (PI3K) and not with changes in the protein levels of the p85 regulatory subunit of PI3K, PDK1 or PTEN- modulators, and/or mediators of PI3K activity. eAIG could be blunted selectively by treatment with the PI3K inhibitor, LY294002, or by overexpression of either the PI3K antagonist, PTEN, dominant-interfering alleles of PI3K or a downstream PI3K mediator, AKT, but not by the MEK inhibitor, PD98059, dominant-interfering alleles of MEK or the signal transducer and activator of transcription (STAT)-3. In contrast, RNAi-mediated knockdown of FAK resulted in increased v-Src-induced AIG. Expression of a constitutively active PI3K allele was sufficient to induce higher levels of AIG, whereas overexpression of v-Src produced only larger-sized colonies in soft agar. Interestingly, FAK was required for full activation of PI3K by PDGF whereas the activation of PI3K by insulin was significantly increased in FAK-/- cells. Thus, although FAK is dispensable for v-Src-induced oncogenic transformation in vitro, it may exert either positive or negative effects on signaling or motility depending on which pathways are activated in cancer cells.
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PMID:Enhanced v-Src-induced oncogenic transformation in the absence of focal adhesion kinase is mediated by phosphatidylinositol 3-kinase. 1580 50

The protein kinase AKT is a key regulator for cell growth, cell survival and metabolic insulin action. However, the mechanism of activation of AKT in vivo, which presumably involves membrane recruitment of the kinase, oligomerization, and multiple phosphorylation events, is not fully understood. In the present study, we have expressed and purified dimeric GST-fusion proteins of human protein kinase AKT2 (DeltaPH-AKT2) in milligram quantities via the baculovirus expression system. Treatment of virus-infected insect cells with the phosphatase inhibitor okadaic acid (OA) led to phosphorylation of the two regulatory phosphorylation sites, Thr309 and Ser474, and to activation of the kinase. Likewise, phosphorylation of Thr309 in vitro by recombinant PDK1 or mutation of Thr309 and Ser474 to acidic residues rendered the kinase constitutively active. However, even though the specific activity of our AKT2 was increased 15-fold compared to previous reports, GST-mediated dimerization alone did not lead to an activation of the kinase. Whereas both mutagenesis and phosphorylation led to an increase in the turnover number of the enzyme, only the latter resulted in a marked reduction (20-fold) of the apparent Km value for the exogenous substrate Crosstide, indicating that this widely used mutagenesis only partially mimics phosphorylation. Kinetic analysis of GST-AKT2 demonstrates that phosphorylation of Thr309 in the activation loop of the kinase is largely responsible for the observed reduction in Km and for a subsequent 150-fold increase in the catalytic efficiency (k(cat)/Km) of the enzyme. Highly active AKT2 constructs were used in autophosphorylation reactions in vitro, where inactive AKT2 kinases served as substrates. As a matter of fact, we found evidence for a minor autophosphorylation activity of AKT2 but no significant autophosphorylation of any of the two regulatory sites, Thr309 or Ser474.
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PMID:Activation of a GST-tagged AKT2/PKBbeta. 1589 Apr 50

Several recent reports have brought conclusive evidence that the tumor suppressor PTEN, once considered a strictly cytoplasmic protein, shuttles to the nuclear compartment, where it joins a variety of components of the same pathway it regulates in the cytoplasm, among which PI3K, PDK1 and AKT. In this review, we focus on the growing supporting evidence for an important physiological role of this nuclear pathway and on the role that alteration of this novel regulatory circuit may play during cell transformation.
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PMID:Class reunion: PTEN joins the nuclear crew. 1628 86

Activation of kinases signalling pathways contributes to various malignant phenotypes in human cancers, including breast tumour. To examine the possible activation of these signalling molecules, we examined the phosphorylation status in 12 protein kinases and transcription factors in normal primary human mammary epithelial cells, telomerase-immortalised human breast epithelial cell line, and two breast cancer lines, MDA-MB-468 and MCF-7, using Kinexus phosphorylated protein screening assays. The phosphorylation of FAK, mTOR, p70S6K, and PDK-1 were elevated in both breast cancer cell lines, whereas the phosphorylation of AKT, EGFR, ErbB2/Her2, PDGFR, Shc, and Stat3 were elevated in only one breast cancer line compared to normal primary mammary epithelial cells and telomerase-immortalised breast epithelial cells. The same findings were confirmed by Western blotting and by kinase assays. We further substantiated the phosphorylation status of these molecules in tissue microarray slides containing 89 invasive breast cancer tissues as well as six normal mammary tissues with immunohistochemistry staining using phospho-specific antibodies. Consistent findings were obtained as greater than 70% of invasive breast carcinomas expressed moderate to high levels of phosphorylated PDK-1, AKT, p70S6K, and EGFR. In sharp contrast, phosphorylation of the same proteins was nearly undetectable or was at low levels in normal mammary tissues under the same assay. Elevated phosphorylation of PDK-1, AKT, mTOR, p70S6K, S6, EGFR, and Stat3 were highly associated with invasive breast tumours (P<0.05). Taken together, our results suggest that activation of these kinase pathways by phosphorylation may in part account for molecular pathogenesis of human breast carcinoma. Particularly, moderate to high level of PDK-1 phosphorylation was found in 86% of high-grade metastasised breast tumours. This is the first report demonstrating phosphorylation of PDK-1 is frequently elevated in breast cancer with concomitantly increased phosphorylation of downstream kinases, including AKT, mTOR, p70S6K, S6, and Stat3. This finding thus suggested PDK-1 may promote oncogenesis in part through the activation of AKT and p70S6K and rationalised that PDK-1 as well as downstream components of PDK-1 signalling pathway may be promising therapeutic targets to treat breast cancer.
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PMID:Elevated phosphorylation and activation of PDK-1/AKT pathway in human breast cancer. 1628 4

ARK5 is a tumor progression-associated factor that is directly phosphorylated by AKT at serine 600 in the regulatory domain, but phosphorylation at the conserved threonine residue on the active T loop has been found to be required for its full activation. In this study, we identified serine/threonine protein kinase NDR2 as a protein kinase that phosphorylates and activates ARK5 during insulin-like growth factor (IGF)-1 signaling. Upon stimulation with IGF-1, NDR2 was found to directly phosphorylate the conserved threonine 211 on the active T loop of ARK5 and to promote cell survival and invasion of colorectal cancer cell lines through ARK5. During IGF-1 signaling, phosphorylation at three residues (threonine 75, serine 282, and threonine 442) was also found to be required for NDR2 activation. Among these three residues, phosphorylation of serine 282 seemed to be the most important for NDR2 activation (the same as for the mouse homologue) because its aspartic acid-converted mutant (NDR2/S282D) induced ARK5-mediated cell survival and invasion activities even in the absence of IGF-1. As in the mouse homologue, threonine 75 in NDR2 was required for interaction with S100B, and binding was in a calcium ion- and phospholipase C-gamma-dependent manner. We also found that PDK-1 plays an important role in NDR2 activation especially in the phosphorylation of threonine 442. Based on the results of this study, we report here that NDR2 is an upstream kinase of ARK5 that plays an essential role in tumor progression through ARK5.
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PMID:NDR2 acts as the upstream kinase of ARK5 during insulin-like growth factor-1 signaling. 1648 89

Phosphoinositol-3-kinase (PI3K)/protein kinase B (AKT) and Fms-like tyrosine kinase 3 (FLT3) signaling are aberrantly activated in acute myelogenous leukemia (AML) cells. Constitutively activated AKT and FLT3 regulate leukemia cell survival and resistance to chemotherapy. In this study, we investigated the effects of the novel multiple kinase inhibitor KP372-1 on the survival of AML cell lines and primary AML samples. KP372-1 directly inhibited the kinase activity of AKT, PDK1, and FLT3 in a concentration-dependent manner. Western blot analysis indicated that KP372-1 decreased the phosphorylation of AKT on both Ser(473) and Thr(308); abrogated the phosphorylation of p70S6 kinase, BAD, and Foxo3a via PI3K/AKT signaling; and down-regulated expression of PIM-1 through direct inhibition of FLT3. Treatment of AML cell lines with KP372-1 resulted in rapid generation of reactive oxygen species and stimulation of oxygen consumption, followed by mitochondrial depolarization, caspase activation, and phosphatidylserine externalization. KP372-1 induced pronounced apoptosis in AML cell lines and primary samples irrespective of their FLT3 status, but not in normal CD34(+) cells. Moreover, KP372-1 markedly decreased the colony-forming ability of primary AML samples (IC(50) < 200 nmol/L) with minimal cytotoxic effects on normal progenitor cells. Taken together, our results show that the simultaneous inhibition of critical prosurvival kinases by KP372-1 leads to mitochondrial dysfunction and apoptosis of AML but not normal hematopoietic progenitor cells.
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PMID:Simultaneous inhibition of PDK1/AKT and Fms-like tyrosine kinase 3 signaling by a small-molecule KP372-1 induces mitochondrial dysfunction and apoptosis in acute myelogenous leukemia. 1658


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