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Sample records for phosphatidylinositol 3-kinase pi3k

  1. Phosphatidylinositol 3-kinase (PI3K) inhibitors as cancer therapeutics

    PubMed Central

    2013-01-01

    Phosphatidylinositol 3-kinases (PI3Ks) are lipid kinases that regulate diverse cellular processes including proliferation, adhesion, survival, and motility. Dysregulated PI3K pathway signaling occurs in one-third of human tumors. Aberrantly activated PI3K signaling also confers sensitivity and resistance to conventional therapies. PI3K has been recognized as an attractive molecular target for novel anti-cancer molecules. In the last few years, several classes of potent and selective small molecule PI3K inhibitors have been developed, and at least fifteen compounds have progressed into clinical trials as new anticancer drugs. Among these, idelalisib has advanced to phase III trials in patients with advanced indolent non-Hodgkin’s lymphoma and mantle cell lymphoma. In this review, we summarized the major molecules of PI3K signaling pathway, and discussed the preclinical models and clinical trials of potent small-molecule PI3K inhibitors. PMID:24261963

  2. Diosgenin inhibits melanogenesis through the activation of phosphatidylinositol-3-kinase pathway (PI3K) signaling.

    PubMed

    Lee, Jongsung; Jung, Kwangseon; Kim, Yeong Shik; Park, Deokhoon

    2007-06-27

    An increased level of melanin is characteristic of a large number of skin diseases, including acquired hyperpigmentation conditions such as melasma, post inflammatory melanoderma, and solar lentigo. Thus, there is an increasing need for the development of depigmenting agents. In order to evaluate the depigmenting capacity of diosgenin and elucidate its mechanism of action, several experiments were performed in B16 melanoma cells. Melanin content and Western blots for proteins that are involved in melanogenesis were assessed in this study. The melanin content was significantly inhibited by diosgenin. To clarify the mechanism of the depigmenting property of diosgenin, we examined the involvement of diosgenin in the phosphatidylinositol-3-kinase (PI3K) pathway. In this study, diosgenin inhibited the reduction of Akt and GSK 3beta phosphorylation induced by LY294,002, a PI3K inhibitor. In accordance with this result, production levels of MITF (microphthalmia-associated transcription factor) and tyrosinase were increased by diosgenin. These data suggest that diosgenin inhibits melanogenesis through the activation of the PI3K pathway. This suggestion was further confirmed by the fact that the increased production level of melanin by LY294,002 was reduced by diosgenin in B16 melanoma cells. Our study shows that diosgenin inhibits melanogenesis by activating the PI3K pathway, and also suggests that diosgenin may be an effective inhibitor of hyperpigmentation.

  3. Putative Phosphatidylinositol 3-Kinase (PI3K) Binding Motifs in Ovine Betaretrovirus Env Proteins Are Not Essential for Rodent Fibroblast Transformation and PI3K/Akt Activation

    PubMed Central

    Liu, Shan-Lu; Lerman, Michael I.; Miller, A. Dusty

    2003-01-01

    Jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus (ENTV) are simple betaretroviruses that cause epithelial cell tumors in the lower and upper airways of sheep and goats. The envelope (Env) glycoproteins of both viruses can transform rodent and chicken fibroblasts, indicating that they play an essential role in oncogenesis. Previous studies found that a YXXM motif in the Env cytoplasmic tail, a putative docking site for phosphatidylinositol 3-kinase (PI3K) after tyrosine phosphorylation, was necessary for rodent cell transformation but was not required for transformation of DF-1 chicken fibroblasts. Here we show that JSRV and ENTV Env proteins with tyrosine or methionine mutations in the YXXM motif can still transform rodent fibroblasts, albeit with reduced efficiency. Akt was activated in cells transformed by JSRV or ENTV Env proteins and in cells transformed by the proteins with tyrosine mutations. Furthermore, the PI3K-specific inhibitor LY294002 could inhibit Akt activation and cell transformation in all cases, indicating that Akt activation and transformation is PI3K dependent. However, we could not detect tyrosine phosphorylation of JSRV or ENTV Env proteins or an interaction between the Env proteins and PI3K in the transformed cells. We found no evidence for mitogen-activated protein kinase activation in cells that were transformed by the JSRV or ENTV Env proteins. We conclude that ovine betaretrovirus Env proteins transform the rodent fibroblasts by indirectly activating the PI3K/Akt pathway. PMID:12829832

  4. Icaritin requires Phosphatidylinositol 3 kinase (PI3K)/Akt signaling to counteract skeletal muscle atrophy following mechanical unloading

    PubMed Central

    ZHANG, Zong-Kang; LI, Jie; LIU, Jin; GUO, Baosheng; LEUNG, Albert; ZHANG, Ge; ZHANG, Bao-Ting

    2016-01-01

    Counteracting muscle atrophy induced by mechanical unloading/inactivity is of great clinical need and challenge. A therapeutic agent that could counteract muscle atrophy following mechanical unloading in safety is desired. This study showed that natural product Icaritin (ICT) could increase the phosphorylation level of Phosphatidylinositol 3 kinase (PI3K) at p110 catalytic subunit and promote PI3K/Akt signaling markers in C2C12 cells. This study further showed that the high dose ICT treatment could significantly attenuate the decreases in the phosphorylation level of PI3K at p110 catalytic subunit and its downstream markers related to protein synthesis, and inhibit the increases in protein degradation markers at mRNA and protein levels in rat soleus muscle following 28-day hindlimb unloading. In addition, the decreases in soleus muscle mass, muscle fiber cross-sectional area, twitch force, specific force, contraction time and half relaxation time could be significantly attenuated by the high dose ICT treatment. The low dose ICT treatment could moderately attenuate the above changes induced by unloading. Wortmannin, a specific inhibitor of PI3K at p110 catalytic subunit, could abolish the above effects of ICT in vitro and in vivo, indicating that PI3K/Akt signaling could be required by ICT to counteract skeletal muscle atrophy following mechanical unloading. PMID:26831566

  5. ErbB3 ablation impairs phosphatidylinositol 3-kinase (PI3K)/AKT-dependent mammary tumorigenesis

    PubMed Central

    Cook, Rebecca S.; Garrett, Joan T.; Sánchez, Violeta; Stanford, Jamie C.; Young, Christian; Chakravarty, Anindita; Rinehart, Cammie; Zhang, Yixian; Wu, Yaming; Greenberger, Lee; Horak, Ivan D.; Arteaga, Carlos L.

    2011-01-01

    Summary The ErbB receptor family member ErbB3 has been implicated in breast cancer growth but it has yet to be determined whether its disruption is therapeutically valuable. In a mouse model of mammary carcinoma driven by the polyomavirus middle T (PyVmT) oncogene, the ErbB2 tyrosine kinase inhibitor lapatinib reduced the activation of ErbB3 and Akt along with tumor cell growth. In this phosphatidylinositol-3 kinase (PI3K)-dependent tumor model, ErbB2 is part of a complex containing PyVmT, p85 (PI3K), ErbB3, and Src, that is disrupted by treatment with lapatinib. Thus, full engagement of PI3K/Akt by ErbB2 in this oncogene-induced mouse tumor model may involve its ability to dimerize with and phosphorylate ErbB3, which itself directly binds PI3K. Here we report that ErbB3 is critical for PI3K/AKT-driven tumor formation triggered by the PyVmT oncogene. Tissue-specific, Cre-mediated deletion of ErbB3 reduced Akt phosphorylation, primary tumor growth and pulmonary metastasis. Further EZN-3920, a chemically stabilized antisense oligonucleotide that targets the ErbB3 mRNA in vivo, produced similar effects while causing no mouse toxicity. Our findings offer further preclinical evidence that ErbB3 ablation may be therapeutically effective in tumors where ErbB3 engages PI3K/Akt signaling. PMID:21482676

  6. Infectious bursal disease virus activates the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway by interaction of VP5 protein with the p85{alpha} subunit of PI3K

    SciTech Connect

    Wei Li; Hou Lei; Zhu Shanshan; Wang Jing; Zhou Jiao; Liu Jue

    2011-08-15

    Phosphatidylinositol 3-kinase (PI3K)/Akt signaling is commonly activated upon virus infection and has been implicated in the regulation of diverse cellular functions such as proliferation and apoptosis. The present study demonstrated for the first time that infectious bursal disease virus (IBDV), the causative agent of a highly contagious disease in chickens, can induce Akt phosphorylation in cultured cells, by a mechanism that is dependent on PI3K. Inhibition of PI3K activation greatly enhanced virus-induced cytopathic effect and apoptotic cell death as evidenced by cleavage of poly-ADP ribose polymerase and activation of caspase-3. Investigations into the mechanism of PI3K/Akt activation revealed that IBDV activates PI3K/Akt signaling through binding of the non-structural protein VP5 to regulatory subunit p85{alpha} of PI3K resulting in the suppression of premature apoptosis and improved virus growth after infection. The results presented here provide a basis for understanding molecular mechanism of IBDV infection.

  7. Phosphatidylinositol 3-Kinase (PI3K) Signaling via Glycogen Synthase Kinase-3 (Gsk-3) Regulates DNA Methylation of Imprinted Loci*

    PubMed Central

    Popkie, Anthony P.; Zeidner, Leigh C.; Albrecht, Ashley M.; D'Ippolito, Anthony; Eckardt, Sigrid; Newsom, David E.; Groden, Joanna; Doble, Bradley W.; Aronow, Bruce; McLaughlin, K. John; White, Peter; Phiel, Christopher J.

    2010-01-01

    Glycogen synthase kinase-3 (Gsk-3) isoforms, Gsk-3α and Gsk-3β, are constitutively active, largely inhibitory kinases involved in signal transduction. Underscoring their biological significance, altered Gsk-3 activity has been implicated in diabetes, Alzheimer disease, schizophrenia, and bipolar disorder. Here, we demonstrate that deletion of both Gsk-3α and Gsk-3β in mouse embryonic stem cells results in reduced expression of the de novo DNA methyltransferase Dnmt3a2, causing misexpression of the imprinted genes Igf2, H19, and Igf2r and hypomethylation of their corresponding imprinted control regions. Treatment of wild-type embryonic stem cells and neural stem cells with the Gsk-3 inhibitor, lithium, phenocopies the DNA hypomethylation at these imprinted loci. We show that inhibition of Gsk-3 by phosphatidylinositol 3-kinase (PI3K)-mediated activation of Akt also results in reduced DNA methylation at these imprinted loci. Finally, we find that N-Myc is a potent Gsk-3-dependent regulator of Dnmt3a2 expression. In summary, we have identified a signal transduction pathway that is capable of altering the DNA methylation of imprinted loci. PMID:21047779

  8. Domain analysis reveals striking functional differences between the regulatory subunits of phosphatidylinositol 3-kinase (PI3K), p85α and p85β.

    PubMed

    Ito, Yoshihiro; Vogt, Peter K; Hart, Jonathan R

    2017-08-22

    Our understanding of isoform-specific activities of phosphatidylinositol 3-kinase (PI3K) is still rudimentary, and yet, deep knowledge of these non-redundant functions in the PI3K family is essential for effective and safe control of PI3K in disease. The two major isoforms of the regulatory subunits of PI3K are p85α and p85β, encoded by the genes PIK3R1 and PIK3R2, respectively. These isoforms show distinct functional differences that affect and control cellular PI3K activity and signaling [1-4]. In this study, we have further explored the differences between p85α and p85β by genetic truncations and substitutions. We have discovered unexpected activities of the mutant proteins that reflect regulatory functions of distinct p85 domains. These results can be summarized as follows: Deletion of the SH3 domain increases oncogenic and PI3K signaling activity. Deletion of the combined SH3-RhoGAP domains abolishes these activities. In p85β, deletion of the cSH2 domain reduces oncogenic and signaling activities. In p85α, such a deletion has an activating effect. The deletions of the combined cSH2 and iSH2 domains and also the deletion of the cSH2, iSH2 and nSH2 domains yield results that go in the same direction, generally activating in p85α and reducing activity in p85β. The contrasting functions of the cSH2 domains are verified by domain exchanges with the cSH2 domain of p85β exerting an activating effect and the cSH2 domain of p85α an inactivating effect, even in the heterologous isoform. In the cell systems studied, protein stability was not correlated with oncogenic and signaling activity. These observations significantly expand our knowledge of the isoform-specific activities of p85α and p85β and of the functional significance of specific domains for regulating the catalytic subunits of class IA PI3K.

  9. Volatile Inhibitors of Phosphatidylinositol-3-Kinase (PI3K) Pathway: Anti-Cancer Potential of Aroma Compounds of Plant Essential Oils.

    PubMed

    Bordoloi, Manobjyoti; Saikia, Surovi; Kolita, Bhaskor; Sarmah, Rajeev

    2017-03-27

    Cancer is a grave health problem for the world as global cancer burden rises to 14 million new cases with 8.2 million deaths every year which is expected to rise by 70% in next 2 decades as reported by WHO. These steady rises in death demand for rapid developments in anti-cancer agents. Essential oils, being natural and multi-component complex systems have recently attracted a lot of attention in this search for novel anti-cancer agents. The pharmaceutical attributes of essential oil components specifically focusing their affinity towards COX, 5-LOX, AKT, MDM2, PDK1 and mTOR which defines the phosphatidylinositol-3-kinase (PI3K) pathway were assessed. 123 compounds present in essential oils of different plants were analyzed for their drug like attributes which were then allowed to dock with PI3K dependent receptors crucial for development of cancer malignancies. Among them, 21 compounds were filtered possessing high druglikeness with favourable metabolism offered by major cytochromeP450 isoforms. Finally, the best docked compounds with highest binding affinities were employed for building a ligand based pharmacophore. Being inhibitors P-glycoprotein they also exhibited good absorption profiles and non-carcinogenic properties. Further from these 21, six compounds were evaluated against A549 lung cancer cells. The pharmacophoric feature obtained can be applied for both designing and screening moieties for active inhibitors of the phosphatidylinositol-3-kinase pathway specifically from essential oil compounds and these final 21 compounds can be further promoted to studies for anti-cancer drug development. Among these six compounds, exhibited promising inhibitory results against A549 lung cancer cells. Further, immunoblotting assay confirmed the efficacy of the compounds for inhibiting mTOR and AKT enzymes which are bandmasters for downstream signaling of the PI3K pathway. Methyl nonanoate, (R)-citronellol, cis-carveol (L-carveol), 3-methyl-Cyclohexanone, 4-carene and

  10. The phosphatidylinositol 3-kinases (PI3K) inhibitor GS-1101 synergistically potentiates histone deacetylase inhibitor-induced proliferation inhibition and apoptosis through the inactivation of PI3K and extracellular signal-regulated kinase pathways.

    PubMed

    Bodo, Juraj; Zhao, Xiaoxian; Sharma, Arishya; Hill, Brian T; Portell, Craig A; Lannutti, Brian J; Almasan, Alexandru; Hsi, Eric D

    2013-10-01

    Previously, we showed that inhibition of the protein kinase C β (PKCβ)/AKT pathway augments engagement of the histone deacetylase inhibitor (HDI)-induced apoptosis in lymphoma cells. In the present study, we investigated the cytotoxicity and mechanisms of cell death induced by the delta isoform-specific phosphatidylinositide 3-kinase (PI3K) inhibitor, GS-1101, in combination with the HDI, panobinostat (LBH589) and suberoylanilide hydroxamic acid (SAHA). Lymphoma cell lines, primary non-Hodgkin Lymphoma (NHL) and chronic lymphocytic leukaemia (CLL) cells were simultaneously treated with the HDI, LBH589 and GS-1101. An interaction of the LBH589/GS-1101 combination was formally examined by using various concentrations of LBH589 and GS-1101. Combined treatment resulted in a synergistic inhibition of proliferation and showed synergistic effect on apoptotic induction in all tested cell lines and primary NHL and CLL cells. This study indicates that interference with PI3K signalling dramatically increases HDI-mediated apoptosis in malignant haematopoietic cells, possibly through both AKT-dependent or AKT- independent mechanisms. Moreover, the increase in HDI-related apoptosis observed in PI3K inhibitor-treated cells appears to be related to the disruption of the extracellular signal-regulated kinase (ERK) signalling pathway. This study provides a strong rational for testing the combination of PI3K inhibitors and HDI in the clinic.

  11. Phosphatidylinositol 3-Kinase (PI3K) Activity Bound to Insulin-like Growth Factor-I (IGF-I) Receptor, which Is Continuously Sustained by IGF-I Stimulation, Is Required for IGF-I-induced Cell Proliferation*

    PubMed Central

    Fukushima, Toshiaki; Nakamura, Yusaku; Yamanaka, Daisuke; Shibano, Takashi; Chida, Kazuhiro; Minami, Shiro; Asano, Tomoichiro; Hakuno, Fumihiko; Takahashi, Shin-Ichiro

    2012-01-01

    Continuous stimulation of cells with insulin-like growth factors (IGFs) in G1 phase is a well established requirement for IGF-induced cell proliferation; however, the molecular components of this prolonged signaling pathway that is essential for cell cycle progression from G1 to S phase are unclear. IGF-I activates IGF-I receptor (IGF-IR) tyrosine kinase, followed by phosphorylation of substrates such as insulin receptor substrates (IRS) leading to binding of signaling molecules containing SH2 domains, including phosphatidylinositol 3-kinase (PI3K) to IRS and activation of the downstream signaling pathways. In this study, we found prolonged (>9 h) association of PI3K with IGF-IR induced by IGF-I stimulation. PI3K activity was present in this complex in thyrocytes and fibroblasts, although tyrosine phosphorylation of IRS was not yet evident after 9 h of IGF-I stimulation. IGF-I withdrawal in mid-G1 phase impaired the association of PI3K with IGF-IR and suppressed DNA synthesis the same as when PI3K inhibitor was added. Furthermore, we demonstrated that Tyr1316-X-X-Met of IGF-IR functioned as a PI3K binding sequence when this tyrosine is phosphorylated. We then analyzed IGF signaling and proliferation of IGF-IR−/− fibroblasts expressing exogenous mutant IGF-IR in which Tyr1316 was substituted with Phe (Y1316F). In these cells, IGF-I stimulation induced tyrosine phosphorylation of IGF-IR and IRS-1/2, but mutated IGF-IR failed to bind PI3K and to induce maximal phosphorylation of GSK3β and cell proliferation in response to IGF-I. Based on these results, we concluded that PI3K activity bound to IGF-IR, which is continuously sustained by IGF-I stimulation, is required for IGF-I-induced cell proliferation. PMID:22767591

  12. CAL-101, a p110δ selective phosphatidylinositol-3-kinase inhibitor for the treatment of B-cell malignancies, inhibits PI3K signaling and cellular viability

    PubMed Central

    Meadows, Sarah A.; Herman, Sarah E. M.; Kashishian, Adam; Steiner, Bart; Johnson, Amy J.; Byrd, John C.; Tyner, Jeffrey W.; Loriaux, Marc M.; Deininger, Mike; Druker, Brian J.; Puri, Kamal D.; Ulrich, Roger G.; Giese, Neill A.

    2011-01-01

    Phosphatidylinositol-3-kinase p110δ serves as a central integration point for signaling from cell surface receptors known to promote malignant B-cell proliferation and survival. This provides a rationale for the development of small molecule inhibitors that selectively target p110δ as a treatment approach for patients with B-cell malignancies. We thus identified 5-fluoro-3-phenyl-2-[(S)-1-(9H-purin-6-ylamino)-propyl]-3H-quinazolin-4-one (CAL-101), a highly selective and potent p110δ small molecule inhibitor (half-maximal effective concentration [EC50] = 8nM). Using tumor cell lines and primary patient samples representing multiple B-cell malignancies, we have demonstrated that constitutive phosphatidylinositol-3-kinase pathway activation is p110δ-dependent. CAL-101 blocked constitutive phosphatidylinositol-3-kinase signaling, resulting in decreased phosphorylation of Akt and other downstream effectors, an increase in poly(ADP-ribose) polymerase and caspase cleavage and an induction of apoptosis. These effects have been observed across a broad range of immature and mature B-cell malignancies, thereby providing a rationale for the ongoing clinical evaluation of CAL-101. PMID:20959606

  13. CAL-101, a p110delta selective phosphatidylinositol-3-kinase inhibitor for the treatment of B-cell malignancies, inhibits PI3K signaling and cellular viability.

    PubMed

    Lannutti, Brian J; Meadows, Sarah A; Herman, Sarah E M; Kashishian, Adam; Steiner, Bart; Johnson, Amy J; Byrd, John C; Tyner, Jeffrey W; Loriaux, Marc M; Deininger, Mike; Druker, Brian J; Puri, Kamal D; Ulrich, Roger G; Giese, Neill A

    2011-01-13

    Phosphatidylinositol-3-kinase p110δ serves as a central integration point for signaling from cell surface receptors known to promote malignant B-cell proliferation and survival. This provides a rationale for the development of small molecule inhibitors that selectively target p110δ as a treatment approach for patients with B-cell malignancies. We thus identified 5-fluoro-3-phenyl-2-[(S)-1-(9H-purin-6-ylamino)-propyl]-3H-quinazolin-4-one (CAL-101), a highly selective and potent p110δ small molecule inhibitor (half-maximal effective concentration [EC(50)] = 8nM). Using tumor cell lines and primary patient samples representing multiple B-cell malignancies, we have demonstrated that constitutive phosphatidylinositol-3-kinase pathway activation is p110δ-dependent. CAL-101 blocked constitutive phosphatidylinositol-3-kinase signaling, resulting in decreased phosphorylation of Akt and other downstream effectors, an increase in poly(ADP-ribose) polymerase and caspase cleavage and an induction of apoptosis. These effects have been observed across a broad range of immature and mature B-cell malignancies, thereby providing a rationale for the ongoing clinical evaluation of CAL-101.

  14. Inhibition of Insulin Signaling in Endothelial Cells by Protein Kinase C-induced Phosphorylation of p85 Subunit of Phosphatidylinositol 3-Kinase (PI3K)*

    PubMed Central

    Maeno, Yasuhiro; Li, Qian; Park, Kyoungmin; Rask-Madsen, Christian; Gao, Benbo; Matsumoto, Motonobu; Liu, Yingjie; Wu, I-Hsien; White, Morris F.; Feener, Edward P.; King, George L.

    2012-01-01

    The regulation of endothelial function by insulin is consistently abnormal in insulin-resistant states and diabetes. Protein kinase C (PKC) activation has been reported to inhibit insulin signaling selectively in endothelial cells via the insulin receptor substrate/PI3K/Akt pathway to reduce the activation of endothelial nitric-oxide synthase (eNOS). In this study, it was observed that PKC activation differentially inhibited insulin receptor substrate 1/2 (IRS1/2) signaling of insulin's activation of PI3K/eNOS by decreasing only tyrosine phosphorylation of IRS2. In addition, PKC activation, by general activator and specifically by angiotensin II, increased the phosphorylation of p85/PI3K, which decreases its association with IRS1 and activation. Thr-86 of p85/PI3K was identified to be phosphorylated by PKC activation and confirmed to affect IRS1-mediated activation of Akt/eNOS by insulin and VEGF using a deletion mutant of the Thr-86 region of p85/PI3K. Thus, PKC and angiotensin-induced phosphorylation of Thr-86 of p85/PI3K may partially inhibit the activation of PI3K/eNOS by multiple cytokines and contribute to endothelial dysfunction in metabolic disorders. PMID:22158866

  15. Phosphatidylinositol-3-kinase (PI3K) gamma plays a central role in blood-brain barrier dysfunction in acute experimental stroke

    PubMed Central

    Jin, Rong; Song, Zifang; Yu, Shiyong; Piazza, Abigail; Nanda, Anil; Penninger, Josef M; Granger, D Neil; Li, Guohong

    2011-01-01

    Background and Purpose Phosphoinositide 3-kinase (PI3K) gamma is linked to inflammation and oxidative stress. This study was conducted to investigate the role of the PI3Kgamma in the blood-brain barrier (BBB) dysfunction and brain damage induced by focal cerebral ischemia/reperfusion. Methods Wild-type and PI3Kgamma knockout mice were subjected to middle cerebral artery occlusion (60 min) followed by reperfusion. Evans blue leakage, brain edema, infarct volumes and neurological deficits were examined. Oxidative stress, neutrophil infiltration, and matrix metallopeptidase-9 (MMP-9) were assessed. Activation of NF-kB and expression of proinflammatory and pro-oxidative genes were studied. Results PI3Kgamma deficiency significantly reduced BBB permeability and brain edema formation, which were time-dependently correlated with preventing the degradation of the tight junction protein, claudin-5, and the basal lamina protein, collagen IV, and the phosphorylation of myosin light chain (MLC) in brain microvessels. PI3Kgamma deficiency suppressed ischemia/reperfusion-induced NF-kB p65 (Ser536) phosphorylation and the expression of the pro-oxidant enzyme NADPH oxidase (Nox1, Nox2, and Nox4) and pro-inflammatory adhesion molecules (E- and P-selectin, ICAM-1) at different time points. These molecular changes were associated with significant inhibition of oxidative stress (superoxide production and malondialdehyde content), neutrophil infiltration, and MMP-9 expression/activity in PI3Kgamma knockout mice. Eventually, PI3Kgamma deficiency significantly reduced infarct volumes and neurological scores at 24 hours after ischemia/reperfusion. Conclusions Our results provide the first direct demonstration that PI3Kgamma plays a significant role in ischemia/reperfusion-induced BBB disruption and brain damage. Future studies need to explore PI3Kγ as a potential target for stroke therapy. PMID:21546487

  16. Intracellular signal transduction pathways in the regulation of fowl sperm motility: evidence for the involvement of phosphatidylinositol 3-kinase (PI3-K) cascade.

    PubMed

    Ashizawa, Koji; Omura, Yusuke; Katayama, Seiichi; Tatemoto, Hideki; Narumi, Kazunori; Tsuzuki, Yasuhiro

    2009-07-01

    The possible role of PI3-K in the reversible temperature-dependent immobilization of fowl sperm motility was investigated by using PI3-K inhibitor (LY294002) and its inactive analogue (LY303511). The existence of the PI3-K in fowl spermatozoa was also confirmed by Western blotting analysis. Fowl sperm motility in TES/NaCl buffer remained negligible at the avian body temperature of 40 degrees C but was maintained vigorously when the temperature was decreased to 30 degrees C. At 30 degrees C, no stimulation or inhibition of motility was observed after the addition of 2 mM CaCl2 and 10 microM LY294002 or LY303511: around 70-80% of spermatozoa remained motile. In contrast, at 40 degrees C, the motility of spermatozoa was activated immediately after the addition of Ca(2+), but the subsequent addition of LY294002 inhibited the motility again. The addition of LY303511 did not appreciably affect the Ca(2+)-supplemented sperm motility, which was maintained for at least 15 min. The ATP concentrations of spermatozoa after the addition of LY294002 + Ca(2+) or LY303511 + Ca(2+) were almost the same values compared with those of Ca(2+) alone at 40 degrees C, suggesting that the addition of LY294002 was not simply affecting membrane damage or inhibiting energy production in the spermatozoa, but may be acting on some part of the motility-regulating cascade. Immunoblotting of sperm extract using an antibody to PI3-K revealed a major cross-reacting protein of 85 kDa, which corresponds to the molecular weight of the subunit of PI3-K. These results suggest that PI3-K may be positively involved in the calcium-regulated maintenance of flagellar movement of fowl spermatozoa at 40 degrees C.

  17. Phosphatidylinositol 3-kinase-AKT-mammalian target of rapamycin (PI3K-Akt-mTOR) signaling pathway in non-small cell lung cancer

    PubMed Central

    2015-01-01

    Non-small cell lung cancer (NSCLC) is a devastating disease with poor prognosis. Systemic chemotherapy has been the mainstay of treatment in advanced disease for many decades. Personalized targeted therapy such as epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) and crizotinib has significantly changed the treatment paradigm in NSCLC. The future success of development of molecular targeted therapy relies on the understanding of signal transduction pathways. The PI3K-Akt-mTOR pathway is commonly deregulated in human malignancy including NSCLC. Therefore, this pathway is a target for many therapeutic developments. This review will provide an overview of PI3K-Akt-mTOR signaling pathway, genetic alterations activating the pathway and clinical therapeutic development of pathway inhibitors. PMID:25870799

  18. Anticarcinogenic action of quercetin by downregulation of phosphatidylinositol 3-kinase (PI3K) and protein kinase C (PKC) via induction of p53 in hepatocellular carcinoma (HepG2) cell line.

    PubMed

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2015-09-01

    Protein kinase C (PKC) is a key regulator of cell growth and differentiation in mammalian cells and hyperactivation of PKC is believed to play an important role in tumor progression. PKC is downstream to signaling protein of phosphatidylinositol 3-Kinase (PI3K), a known up-regulator of cell proliferation and survival. Accumulation of reactive oxygen species (ROS) triggers oxidative stress in the tumor microenvironment, leading to the hyperactivation of various oxidative stress-stimulated signaling molecules. Quercetin (QUE) is a naturally occurring dietary flavonoid having antioxidant properties. QUE is reported to show antitumor activity both in vitro and in vivo; however, the molecular mechanism is yet to be thoroughly explored. HepG2 cells display cellular functions similar to the normal hepatocytes with high degree of morphological and functional differentiation, therefore HepG2 cell line is chosen as the suitable model for drug targeting. Present study is aimed to establish the signaling pathway involved in the anticarcinogenic action of QUE in HepG2 cell line. HepG2 cells were treated with different doses of QUE. Protein level and gene expression were analysed by Western blotting and RT-PCR, respectively. PKC activity was measured by non-radioactive-tagged phosphorylation. Results showed downregulation of expression of PI3K, PKC, COX-2 and ROS caused by QUE. Additionally, QUE enhanced the expression of p53 and BAX in HepG2 cells. Overall, results of the current study suggested that QUE elicited anticarcinogenic action by upregulation of p53 and BAX in HepG2 cells via downregulation of ROS, PKC, PI3K and COX-2, confirming our earlier report on the animal model.

  19. A Phase Ib Study of BEZ235, a Dual Inhibitor of Phosphatidylinositol 3-Kinase (PI3K) and Mammalian Target of Rapamycin (mTOR), in Patients With Advanced Renal Cell Carcinoma

    PubMed Central

    Molina, Ana M.; Lakhman, Yulia; Patil, Sujata; Woo, Kaitlin; DeLuca, John; Lee, Chung-Han; Hsieh, James J.; Feldman, Darren R.; Motzer, Robert J.; H. Voss, Martin

    2016-01-01

    Lessons Learned Our results highlight additional toxicities of dual PI3K/mTOR inhibition in the clinical setting that were unforeseen from preclinical models. Because of toxicity and lack of efficacy, BEZ235 should not be further developed in the current formulation for patients with renal cell carcinoma. Background. Allosteric inhibitors of the mammalian target of rapamycin complex 1 (mTORC1) are approved for advanced renal cell carcinoma (RCC). Preclinical models have suggested that dual inhibition of phosphatidylinositol 3-kinase (PI3K) and mTOR kinase may establish superior anticancer effect. We aimed to establish safety for BEZ235, a potent inhibitor of both PI3K and mTOR, in advanced RCC. Methods. Patients with advanced RCC who had previously failed standard therapy received escalating doses of BEZ235 in sachet formulation twice daily until progression or unacceptable toxicity. Primary endpoints were to identify the maximally tolerated dose (MTD) and to determine the recommended dose for the phase II study. Results. The study was terminated early because of high incidence of dose-limiting toxicities (DLTs) across all dose levels tested. Ten patients were treated with BEZ235—six with clear cell and four with non-clear cell subtypes. Five of these patients suffered DLTs: 2 of 2 patients in the original 400 mg b.i.d. cohort, 1 of 6 in the 200 mg b.i.d. cohort, and 2 of 2 in the 300 mg b.i.d. cohort. DLTs included fatigue, rash, nausea and vomiting, diarrhea, mucositis, anorexia, and dysgeusia. Five patients were evaluable for response: Two had stable disease as best response, and three had progressive disease. Conclusion. BEZ235 twice daily resulted in significant toxicity without objective responses; further development of this compound will not be pursued in this disease. PMID:27286790

  20. E6 variants of human papillomavirus 18 differentially modulate the protein kinase B/phosphatidylinositol 3-kinase (akt/PI3K) signaling pathway

    SciTech Connect

    Contreras-Paredes, Adriana

    2009-01-05

    Intra-type genome variations of high risk Human papillomavirus (HPV) have been associated with a differential threat for cervical cancer development. In this work, the effect of HPV18 E6 isolates in Akt/PKB and Mitogen-associated protein kinase (MAPKs) signaling pathways and its implication in cell proliferation were analyzed. E6 from HPV types 16 and 18 are able to bind and promote degradation of Human disc large (hDlg). Our results show that E6 variants differentially modulate hDlg degradation, rebounding in levels of activated PTEN and PKB. HPV18 E6 variants are also able to upregulate phospho-PI3K protein, strongly correlating with activated MAPKs and cell proliferation. Data was supported by the effect of E6 silencing in HPV18-containing HeLa cells, as well as hDlg silencing in the tested cells. Results suggest that HPV18 intra-type variations may derive in differential abilities to activate cell-signaling pathways such as Akt/PKB and MAPKs, directly involved in cell survival and proliferation.

  1. Phosphatidylinositol 3-Kinase Class II α-Isoform PI3K-C2α Is Required for Transforming Growth Factor β-induced Smad Signaling in Endothelial Cells*

    PubMed Central

    Aki, Sho; Yoshioka, Kazuaki; Okamoto, Yasuo; Takuwa, Noriko; Takuwa, Yoh

    2015-01-01

    We have recently demonstrated that the PI3K class II-α isoform (PI3K-C2α), which generates phosphatidylinositol 3-phosphate and phosphatidylinositol 3,4-bisphosphates, plays crucial roles in angiogenesis, by analyzing PI3K-C2α knock-out mice. The PI3K-C2α actions are mediated at least in part through its participation in the internalization of VEGF receptor-2 and sphingosine-1-phosphate receptor S1P1 and thereby their signaling on endosomes. TGFβ, which is also an essential angiogenic factor, signals via the serine/threonine kinase receptor complex to induce phosphorylation of Smad2 and Smad3 (Smad2/3). SARA (Smad anchor for receptor activation) protein, which is localized in early endosomes through its FYVE domain, is required for Smad2/3 signaling. In the present study, we showed that PI3K-C2α knockdown nearly completely abolished TGFβ1-induced phosphorylation and nuclear translocation of Smad2/3 in vascular endothelial cells (ECs). PI3K-C2α was necessary for TGFβ-induced increase in phosphatidylinositol 3,4-bisphosphates in the plasma membrane and TGFβ receptor internalization into the SARA-containing early endosomes, but not for phosphatidylinositol 3-phosphate enrichment or localization of SARA in the early endosomes. PI3K-C2α was also required for TGFβ receptor-mediated formation of SARA-Smad2/3 complex. Inhibition of dynamin, which is required for the clathrin-dependent receptor endocytosis, suppressed both TGFβ receptor internalization and Smad2/3 phosphorylation. TGFβ1 stimulated Smad-dependent VEGF-A expression, VEGF receptor-mediated EC migration, and capillary-like tube formation, which were all abolished by either PI3K-C2α knockdown or a dynamin inhibitor. Finally, TGFβ1-induced microvessel formation in Matrigel plugs was greatly attenuated in EC-specific PI3K-C2α-deleted mice. These observations indicate that PI3K-C2α plays the pivotal role in TGFβ receptor endocytosis and thereby Smad2/3 signaling, participating in angiogenic

  2. First-in-human Phase I study of Pictilisib (GDC-0941), a potent pan-class I phosphatidylinositol-3-kinase (PI3K) inhibitor, in patients with advanced solid tumors

    PubMed Central

    Baird, Richard; Kristeleit, Rebecca; Shah, Krunal; Moreno, Victor; Clarke, Paul A.; Raynaud, Florence I.; Levy, Gallia; Ware, Joseph A; Mazina, Kathryn; Lin, Ray; Wu, Jenny; Fredrickson, Jill; Spoerke, Jill M; Lackner, Mark R; Yan, Yibing; Friedman, Lori S.; Kaye, Stan B.; Derynck, Mika K.; Workman, Paul; de Bono, Johann S.

    2014-01-01

    Purpose This first-in-human dose-escalation trial evaluated the safety, tolerability, maximal tolerated dose (MTD), dose limiting toxicities (DLTs), pharmacokinetics, pharmacodynamics and preliminary clinical activity of pictilisib (GDC-0941), an oral, potent and selective inhibitor of the Class I phosphatidylinositol-3-kinases (PI3K). Patients and Methods Sixty patients with solid tumors received pictilisib at 14 dose levels from 15 to 450mg once-daily, initially on days 1-21 every 28 days and later, utilizing continuous dosing for selected dose levels. Pharmacodynamic studies incorporated 18F-FDG-PET, and assessment of phosphorylated AKT and S6 ribosomal protein in platelet-rich plasma and tumor tissue. Results Pictilisib was well-tolerated. The most common toxicities were grade 1-2 nausea, rash and fatigue while the DLT was grade 3 maculopapular rash (450mg, 2 of 3 patients; 330mg, 1 of 7 patients). The pharmacokinetic profile was dose-proportional and supported once-daily dosing. Levels of phosphorylated serine-473 AKT were suppressed >90% in platelet rich plasma at 3 hours post-dose at the MTD and in tumor at pictilisib doses associated with AUC >20uM.hr. Significant increase in plasma insulin and glucose levels, and >25% decrease in 18F-FDG uptake by PET in 7 of 32 evaluable patients confirmed target modulation. A patient with V600E BRAF mutant melanoma and another with platinum-refractory epithelial ovarian cancer exhibiting PTEN loss and PIK3CA amplification demonstrated partial response by RECIST and GCIG-CA125 criteria, respectively. Conclusion Pictilisib was safely administered with a dose-proportional pharmacokinetic profile, on-target pharmacodynamic activity at dose levels ≥100mg and signs of antitumor activity. The recommended Phase II dose was continuous dosing at 330mg once-daily. PMID:25370471

  3. First-in-human phase I study of pictilisib (GDC-0941), a potent pan-class I phosphatidylinositol-3-kinase (PI3K) inhibitor, in patients with advanced solid tumors.

    PubMed

    Sarker, Debashis; Ang, Joo Ern; Baird, Richard; Kristeleit, Rebecca; Shah, Krunal; Moreno, Victor; Clarke, Paul A; Raynaud, Florence I; Levy, Gallia; Ware, Joseph A; Mazina, Kathryn; Lin, Ray; Wu, Jenny; Fredrickson, Jill; Spoerke, Jill M; Lackner, Mark R; Yan, Yibing; Friedman, Lori S; Kaye, Stan B; Derynck, Mika K; Workman, Paul; de Bono, Johann S

    2015-01-01

    This first-in-human dose-escalation trial evaluated the safety, tolerability, maximal-tolerated dose (MTD), dose-limiting toxicities (DLT), pharmacokinetics, pharmacodynamics, and preliminary clinical activity of pictilisib (GDC-0941), an oral, potent, and selective inhibitor of the class I phosphatidylinositol-3-kinases (PI3K). Sixty patients with solid tumors received pictilisib at 14 dose levels from 15 to 450 mg once-daily, initially on days 1 to 21 every 28 days and later, using continuous dosing for selected dose levels. Pharmacodynamic studies incorporated (18)F-FDG-PET, and assessment of phosphorylated AKT and S6 ribosomal protein in platelet-rich plasma (PRP) and tumor tissue. Pictilisib was well tolerated. The most common toxicities were grade 1-2 nausea, rash, and fatigue, whereas the DLT was grade 3 maculopapular rash (450 mg, 2 of 3 patients; 330 mg, 1 of 7 patients). The pharmacokinetic profile was dose-proportional and supported once-daily dosing. Levels of phosphorylated serine-473 AKT were suppressed >90% in PRP at 3 hours after dose at the MTD and in tumor at pictilisib doses associated with AUC >20 h·μmol/L. Significant increase in plasma insulin and glucose levels, and >25% decrease in (18)F-FDG uptake by PET in 7 of 32 evaluable patients confirmed target modulation. A patient with V600E BRAF-mutant melanoma and another with platinum-refractory epithelial ovarian cancer exhibiting PTEN loss and PIK3CA amplification demonstrated partial response by RECIST and GCIG-CA125 criteria, respectively. Pictilisib was safely administered with a dose-proportional pharmacokinetic profile, on-target pharmacodynamic activity at dose levels ≥100 mg and signs of antitumor activity. The recommended phase II dose was continuous dosing at 330 mg once-daily. ©2014 American Association for Cancer Research.

  4. The Phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3) Binder Rasa3 Regulates Phosphoinositide 3-kinase (PI3K)-dependent Integrin αIIbβ3 Outside-in Signaling*

    PubMed Central

    Battram, Anthony M.; Durrant, Tom N.; Agbani, Ejaife O.; Heesom, Kate J.; Paul, David S.; Piatt, Raymond; Poole, Alastair W.; Cullen, Peter J.; Bergmeier, Wolfgang; Moore, Samantha F.; Hers, Ingeborg

    2017-01-01

    The class I PI3K family of lipid kinases plays an important role in integrin αIIbβ3 function, thereby supporting thrombus growth and consolidation. Here, we identify Ras/Rap1GAP Rasa3 (GAP1IP4BP) as a major phosphatidylinositol 3,4,5-trisphosphate-binding protein in human platelets and a key regulator of integrin αIIbβ3 outside-in signaling. We demonstrate that cytosolic Rasa3 translocates to the plasma membrane in a PI3K-dependent manner upon activation of human platelets. Expression of wild-type Rasa3 in integrin αIIbβ3-expressing CHO cells blocked Rap1 activity and integrin αIIbβ3-mediated spreading on fibrinogen. In contrast, Rap1GAP-deficient (P489V) and Ras/Rap1GAP-deficient (R371Q) Rasa3 had no effect. We furthermore show that two Rasa3 mutants (H794L and G125V), which are expressed in different mouse models of thrombocytopenia, lack both Ras and Rap1GAP activity and do not affect integrin αIIbβ3-mediated spreading of CHO cells on fibrinogen. Platelets from thrombocytopenic mice expressing GAP-deficient Rasa3 (H794L) show increased spreading on fibrinogen, which in contrast to wild-type platelets is insensitive to PI3K inhibitors. Together, these results support an important role for Rasa3 in PI3K-dependent integrin αIIbβ3-mediated outside-in signaling and cell spreading. PMID:27903653

  5. G protein-coupled receptors (GPCRs) That Signal via Protein Kinase A (PKA) Cross-talk at Insulin Receptor Substrate 1 (IRS1) to Activate the phosphatidylinositol 3-kinase (PI3K)/AKT Pathway.

    PubMed

    Law, Nathan C; White, Morris F; Hunzicker-Dunn, Mary E

    2016-12-30

    G protein-coupled receptors (GPCRs) activate PI3K/v-AKT thymoma viral oncoprotein (AKT) to regulate many cellular functions that promote cell survival, proliferation, and growth. However, the mechanism by which GPCRs activate PI3K/AKT remains poorly understood. We used ovarian preantral granulosa cells (GCs) to elucidate the mechanism by which the GPCR agonist FSH via PKA activates the PI3K/AKT cascade. Insulin-like growth factor 1 (IGF1) is secreted in an autocrine/paracrine manner by GCs and activates the IGF1 receptor (IGF1R) but, in the absence of FSH, fails to stimulate YXXM phosphorylation of IRS1 (insulin receptor substrate 1) required for PI3K/AKT activation. We show that PKA directly phosphorylates the protein phosphatase 1 (PP1) regulatory subunit myosin phosphatase targeting subunit 1 (MYPT1) to activate PP1 associated with the IGF1R-IRS1 complex. Activated PP1 is sufficient to dephosphorylate at least four IRS1 Ser residues, Ser(318), Ser(346), Ser(612), and Ser(789), and promotes IRS1 YXXM phosphorylation by the IGF1R to activate the PI3K/AKT cascade. Additional experiments indicate that this mechanism also occurs in breast cancer, thyroid, and preovulatory granulosa cells, suggesting that the PKA-dependent dephosphorylation of IRS1 Ser/Thr residues is a conserved mechanism by which GPCRs signal to activate the PI3K/AKT pathway downstream of the IGF1R.

  6. Signaling through the Phosphatidylinositol 3-Kinase (PI3K)/Mammalian Target of Rapamycin (mTOR) Axis Is Responsible for Aerobic Glycolysis mediated by Glucose Transporter in Epidermal Growth Factor Receptor (EGFR)-mutated Lung Adenocarcinoma.

    PubMed

    Makinoshima, Hideki; Takita, Masahiro; Saruwatari, Koichi; Umemura, Shigeki; Obata, Yuuki; Ishii, Genichiro; Matsumoto, Shingo; Sugiyama, Eri; Ochiai, Atsushi; Abe, Ryo; Goto, Koichi; Esumi, Hiroyasu; Tsuchihara, Katsuya

    2015-07-10

    Oncogenic epidermal growth factor receptor (EGFR) signaling plays an important role in regulating global metabolic pathways, including aerobic glycolysis, the pentose phosphate pathway (PPP), and pyrimidine biosynthesis. However, the molecular mechanism by which EGFR signaling regulates cancer cell metabolism is still unclear. To elucidate how EGFR signaling is linked to metabolic activity, we investigated the involvement of the RAS/MEK/ERK and PI3K/AKT/mammalian target of rapamycin (mTOR) pathways on metabolic alteration in lung adenocarcinoma (LAD) cell lines with activating EGFR mutations. Although MEK inhibition did not alter lactate production and the extracellular acidification rate, PI3K/mTOR inhibitors significantly suppressed glycolysis in EGFR-mutant LAD cells. Moreover, a comprehensive metabolomics analysis revealed that the levels of glucose 6-phosphate and 6-phosphogluconate as early metabolites in glycolysis and PPP were decreased after inhibition of the PI3K/AKT/mTOR pathway, suggesting a link between PI3K signaling and the proper function of glucose transporters or hexokinases in glycolysis. Indeed, PI3K/mTOR inhibition effectively suppressed membrane localization of facilitative glucose transporter 1 (GLUT1), which, instead, accumulated in the cytoplasm. Finally, aerobic glycolysis and cell proliferation were down-regulated when GLUT1 gene expression was suppressed by RNAi. Taken together, these results suggest that PI3K/AKT/mTOR signaling is indispensable for the regulation of aerobic glycolysis in EGFR-mutated LAD cells. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Small molecule inhibitors of phosphoinositide 3-kinase (PI3K) delta and gamma.

    PubMed

    Ameriks, Michael K; Venable, Jennifer D

    2009-01-01

    In recent years, pharmaceutical companies have increasingly focused on phosphoinositide 3-kinases delta (PI3Kdelta) and gamma (PI3Kgamma) as therapeutic targets for the treatment of inflammatory and autoimmune diseases. All class 1 PI3-kinases (alpha/beta/gamma/delta) generate phospholipid second messengers that help govern cellular processes such as migration, proliferation, and apoptosis. PI3K delta/ gamma lipid kinases are mainly restricted to the hematopoetic system whereas PI3K alpha/beta are ubiquitously expressed, thus raising potential toxicity concerns for chronic indications such as asthma and rheumatoid arthritis. Therefore, the challenge in developing a small molecule inhibitor of PI3K is to define and attain the appropriate isoform selectivity profile. Significant advances in the design of such compounds have been achieved by utilizing x-ray crystal structures of various inhibitors bound to PI3Kgamma in conjunction with pharmacophore modeling and high-throughput screening. Herein, we review the history and challenges involved with the discovery of small molecule isoform-specific PI3K inhibitors. Recent progress in the design of selective PI3Kdelta, PI3Kgamma, and PI3Kdelta/gamma dual inhibitors will be presented.

  8. Phosphatidylinositide 3-kinase (PI3K) and PI3K-related kinase (PIKK) activity contributes to radioresistance in thyroid carcinomas

    PubMed Central

    Burrows, Natalie; Williams, Joseph; Telfer, Brian A; Resch, Julia; Valentine, Helen R; Fitzmaurice, Richard J; Eustace, Amanda; Irlam, Joely; Rowling, Emily J; Hoang-Vu, Cuong; West, Catharine M; Brabant, Georg; Williams, Kaye J

    2016-01-01

    Anaplastic (ATC) and certain follicular thyroid-carcinomas (FTCs) are radioresistant. The Phosphatidylinositide 3-kinase (PI3K) pathway is commonly hyperactivated in thyroid-carcinomas. PI3K can modify the PI3K-related kinases (PIKKs) in response to radiation: How PIKKs interact with PI3K and contribute to radioresistance in thyroid-carcinomas is unknown. Further uncertainties exist in how these interactions function under the radioresistant hypoxic microenvironment. Under normoxia/anoxia, ATC (8505c) and FTC (FTC-133) cells were irradiated, with PI3K-inhibition (via GDC-0941 and PTEN-reconstitution into PTEN-null FTC-133s) and effects on PIKK-activation, DNA-damage, clonogenic-survival and cell cycle, assessed. FTC-xenografts were treated with 5 × 2 Gy, ± 50 mg/kg GDC-0941 (twice-daily; orally) for 14 days and PIKK-activation and tumour-growth assessed. PIKK-expression was additionally assessed in 12 human papillary thyroid-carcinomas, 13 FTCs and 12 ATCs. GDC-0941 inhibited radiation-induced activation of Ataxia-telangiectasia mutated (ATM), ATM-and Rad3-related (ATR) and DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Inhibition of ATM and DNA-PKcs was PI3K-dependent, since activation was reduced in PTEN-reconstituted FTC-133s. Inhibition of PIKK-activation was greater under anoxia: Consequently, whilst DNA-damage was increased and prolonged under both normoxia and anoxia, PI3K-inhibition only reduced clonogenic-survival under anoxia. GDC-0941 abrogated radiation-induced cell cycle arrest, an effect most likely linked to the marked inhibition of ATR-activation. Importantly, GDC-0941 inhibited radiation-induced PIKK-activation in FTC-xenografts leading to a significant increase in time taken for tumours to triple in size: 26.5 ± 5 days (radiation-alone) versus 31.5 ± 5 days (dual-treatment). PIKKs were highly expressed across human thyroid-carcinoma classifications, with ATM scoring consistently lower. Interestingly, some loss of ATM and DNA

  9. Real-time imaging reveals that P2Y2 and P2Y12 receptor agonists are not chemoattractants and macrophage chemotaxis to complement C5a is phosphatidylinositol 3-kinase (PI3K)- and p38 mitogen-activated protein kinase (MAPK)-independent.

    PubMed

    Isfort, Katrin; Ebert, Franziska; Bornhorst, Julia; Sargin, Sarah; Kardakaris, Rozina; Pasparakis, Manolis; Bähler, Martin; Schwerdtle, Tanja; Schwab, Albrecht; Hanley, Peter J

    2011-12-30

    Adenosine 5'-triphosphate (ATP) has been implicated in the recruitment of professional phagocytes (neutrophils and macrophages) to sites of infection and tissue injury in two distinct ways. First, ATP itself is thought to be a chemotactic "find me" signal released by dying cells, and second, autocrine ATP signaling is implicated as an amplifier mechanism for chemotactic navigation to end-target chemoattractants, such as complement C5a. Here we show using real-time chemotaxis assays that mouse peritoneal macrophages do not directionally migrate to stable analogs of ATP (adenosine-5'-(γ-thio)-triphosphate (ATPγS)) or its hydrolysis product ADP (adenosine-5'-(β-thio)-diphosphate (ADPβS)). HPLC revealed that these synthetic P2Y(2) (ATPγS) and P2Y(12) (ADPβS) receptor ligands were in fact slowly degraded. We also found that ATPγS, but not ADPβS, promoted chemokinesis (increased random migration). Furthermore, we found that photorelease of ATP or ADP induced lamellipodial membrane extensions. At the cell signaling level, C5a, but not ATPγS, activated Akt, whereas both ligands induced p38 MAPK activation. p38 MAPK and Akt activation are strongly implicated in neutrophil chemotaxis. However, we found that inhibitors of phosphatidylinositol 3-kinase (PI3K; upstream of Akt) and p38 MAPK (or conditional deletion of p38α MAPK) did not impair macrophage chemotactic efficiency or migration velocity. Our results suggest that PI3K and p38 MAPK are redundant for macrophage chemotaxis and that purinergic P2Y(2) and P2Y(12) receptor ligands are not chemotactic. We propose that ATP signaling is strictly autocrine or paracrine and that ATP and ADP may act as short-range "touch me" (rather than long-range find me) signals to promote phagocytic clearance via cell spreading.

  10. Real-time Imaging Reveals That P2Y2 and P2Y12 Receptor Agonists Are Not Chemoattractants and Macrophage Chemotaxis to Complement C5a Is Phosphatidylinositol 3-Kinase (PI3K)- and p38 Mitogen-activated Protein Kinase (MAPK)-independent*♦

    PubMed Central

    Isfort, Katrin; Ebert, Franziska; Bornhorst, Julia; Sargin, Sarah; Kardakaris, Rozina; Pasparakis, Manolis; Bähler, Martin; Schwerdtle, Tanja; Schwab, Albrecht; Hanley, Peter J.

    2011-01-01

    Adenosine 5′-triphosphate (ATP) has been implicated in the recruitment of professional phagocytes (neutrophils and macrophages) to sites of infection and tissue injury in two distinct ways. First, ATP itself is thought to be a chemotactic “find me” signal released by dying cells, and second, autocrine ATP signaling is implicated as an amplifier mechanism for chemotactic navigation to end-target chemoattractants, such as complement C5a. Here we show using real-time chemotaxis assays that mouse peritoneal macrophages do not directionally migrate to stable analogs of ATP (adenosine-5′-(γ-thio)-triphosphate (ATPγS)) or its hydrolysis product ADP (adenosine-5′-(β-thio)-diphosphate (ADPβS)). HPLC revealed that these synthetic P2Y2 (ATPγS) and P2Y12 (ADPβS) receptor ligands were in fact slowly degraded. We also found that ATPγS, but not ADPβS, promoted chemokinesis (increased random migration). Furthermore, we found that photorelease of ATP or ADP induced lamellipodial membrane extensions. At the cell signaling level, C5a, but not ATPγS, activated Akt, whereas both ligands induced p38 MAPK activation. p38 MAPK and Akt activation are strongly implicated in neutrophil chemotaxis. However, we found that inhibitors of phosphatidylinositol 3-kinase (PI3K; upstream of Akt) and p38 MAPK (or conditional deletion of p38α MAPK) did not impair macrophage chemotactic efficiency or migration velocity. Our results suggest that PI3K and p38 MAPK are redundant for macrophage chemotaxis and that purinergic P2Y2 and P2Y12 receptor ligands are not chemotactic. We propose that ATP signaling is strictly autocrine or paracrine and that ATP and ADP may act as short-range “touch me” (rather than long-range find me) signals to promote phagocytic clearance via cell spreading. PMID:22057273

  11. Epicatechin induces NF-kappaB, activator protein-1 (AP-1) and nuclear transcription factor erythroid 2p45-related factor-2 (Nrf2) via phosphatidylinositol-3-kinase/protein kinase B (PI3K/AKT) and extracellular regulated kinase (ERK) signalling in HepG2 cells.

    PubMed

    Granado-Serrano, Ana Belén; Martín, María Angeles; Haegeman, Guy; Goya, Luis; Bravo, Laura; Ramos, Sonia

    2010-01-01

    The dietary flavonoid epicatechin has been reported to exhibit a wide range of biological activities. The objective of the present study was to investigate the time-dependent regulation by epicatechin on the activity of the main transcription factors (NF-kappaB, activator protein-1 (AP-1) and nuclear transcription factor erythroid 2p45-related factor (Nrf2)) related to antioxidant defence and survival and proliferation pathways in HepG2 cells. Treatment of cells with 10 microm-epicatechin induced the NF-kappaB pathway in a time-dependent manner characterised by increased levels of IkappaB kinase (IKK) and phosphorylated inhibitor of kappaB subunit-alpha (p-IkappaBalpha) and proteolytic degradation of IkappaB, which was consistent with an up-regulation of the NF-kappaB-binding activity. Time-dependent activation of the AP-1 pathway, in concert with enhanced c-Jun nuclear levels and induction of Nrf2 translocation and phosphorylation were also demonstrated. Additionally, epicatechin-induced NF-kappaB and Nrf2 were connected to reactive oxygen species intracellular levels and to the activation of cell survival and proliferation pathways, being phosphatidylinositol-3-kinase/protein kinase B (PI3K/AKT) and extracellular regulated kinase (ERK) associated to Nrf2 modulation and ERK to NF-kappaB induction. These data suggest that the epicatechin-induced survival effect occurs by the induction of redox-sensitive transcription factors through a tight regulation of survival and proliferation pathways.

  12. Clinical development of phosphatidylinositol 3-kinase inhibitors for cancer treatment

    PubMed Central

    2012-01-01

    The phosphatidylinositol 3-kinase (PI3K) pathway is commonly deregulated in cancer. In recent years, the results of the first phase I clinical trials with PI3K inhibitors have become available. In comparison to other targeted agents such v-raf murine sarcoma viral oncogene homolog B1 (BRAF) inhibitors in melanoma or crizotinib in anaplastic lymphoma receptor tyrosine kinase (ALK) translocated tumors, the number of objective responses to PI3K inhibitors is less dramatic. In this review we propose possible strategies to optimize the clinical development of PI3K inhibitors: by exploring the potential role of PI3K isoform-specific inhibitors in improving the therapeutic index, molecular characterization as a basis for patient selection, and the relevance of performing serial tumor biopsies to understand the associated mechanisms of drug resistance. The main focus of this review will be on PI3K isoform-specific inhibitors by describing the functions of different PI3K isoforms, the preclinical activity of selective PI3K isoform-specific inhibitors and the early clinical data of these compounds. PMID:23232172

  13. Drug-resistant phosphatidylinositol 3-kinase: guidance for the preemptive strike.

    PubMed

    Vogt, Peter K

    2008-08-12

    In this issue of Cancer Cell, Zunder et al. (2008) describe surprising findings from investigating inhibitor-resistant mutations in the affinity pocket of p110 alpha of phosphatidylinositol 3-kinase (PI3K). Information on these critical residues provides a road map for generating novel PI3K inhibitors that can overcome the anticipated resistance mutations.

  14. Phosphatidylinositol 3-Kinase and Antiestrogen Resistance in Breast Cancer

    PubMed Central

    Miller, Todd W.; Balko, Justin M.; Arteaga, Carlos L.

    2011-01-01

    Although antiestrogen therapies targeting estrogen receptor (ER) α signaling prevent disease recurrence in the majority of patients with hormone-dependent breast cancer, a significant fraction of patients exhibit de novo or acquired resistance. Currently, the only accepted mechanism linked with endocrine resistance is amplification or overexpression of the ERBB2 (human epidermal growth factor receptor 2 [HER2]) proto-oncogene. Experimental and clinical evidence suggests that hyperactivation of the phosphatidylinositol 3-kinase (PI3K) pathway, the most frequently mutated pathway in breast cancer, promotes antiestrogen resistance. PI3K is a major signaling hub downstream of HER2 and other receptor tyrosine kinases. PI3K activates several molecules involved in cell-cycle progression and survival, and in ER-positive breast cancer cells, it promotes estrogen-dependent and -independent ER transcriptional activity. Preclinical tumor models of antiestrogen-resistant breast cancer often remain sensitive to estrogens and PI3K inhibition, suggesting that simultaneous targeting of the PI3K and ER pathways may be most effective. Herein, we review alterations in the PI3K pathway associated with resistance to endocrine therapy, the state of clinical development of PI3K inhibitors, and strategies for the clinical investigation of such drugs in hormone receptor–positive breast cancer. PMID:22010023

  15. Investigation into the Role of PI3K and JAK3 Kinase Inhibitors in Murine Models of Asthma

    PubMed Central

    Wagh, Akshaya D.; Sharma, Manoranjan; Mahapatra, Jogeshwar; Chatterjee, Abhijeet; Jain, Mukul; Addepalli, Veeranjaneyulu

    2017-01-01

    Asthma is a clinical disorder commonly characterized by chronic eosinophilic inflammation, remodeling and hyper responsiveness of the airways. However, the kinases like Phosphoinositide 3 kinase (PI3K) and Janus kinase 3 (JAK3) are involved in mast cell proliferation, activation, recruitment, migration, and prolonged survival of inflammatory cells. The present study was designed to evaluate the in-vivo comparative effects of two kinase inhibitors on airway inflammation and airway remodeling in acute and chronic models of asthma. Mice were sensitized twice intra-peritoneally and then challenged by inhalation with ovalbumin (OVA). They developed an extensive inflammatory response, goblet cell hyperplasia, collagen deposition, airway smooth muscle thickening similar to pathologies observed in human asthma. The effects of PI3K inhibitor (30 mg/kg, p.o), JAK3 inhibitor (30 mg/kg, p.o) and Dexamethasone (0.3 mg/kg) on airway inflammation and remodeling in OVA sensitized/challenged BALB/c mice were evaluated. Twenty-four hours after the final antigen challenge, bronchoalveolar lavage (BAL) and histological examinations were carried out. It was observed that kinase inhibitors significantly reduced airway inflammation as evidenced by the decrease in pro inflammatory cytokines in BALF and lung homogenate and inflammatory cell count in sensitized mice after allergen challenge. Lung histological analysis showed increased infiltration of inflammatory cells, hyperplasia of goblet cells and the collagen deposition, which were significantly reduced with kinase inhibitor. In conclusion, our data suggest that PI3K and JAK3 inhibitors showed promising alternative therapeutic activity in asthma, which might significantly counteract the airway inflammation in patients with allergic asthma. PMID:28293189

  16. Investigation into the Role of PI3K and JAK3 Kinase Inhibitors in Murine Models of Asthma.

    PubMed

    Wagh, Akshaya D; Sharma, Manoranjan; Mahapatra, Jogeshwar; Chatterjee, Abhijeet; Jain, Mukul; Addepalli, Veeranjaneyulu

    2017-01-01

    Asthma is a clinical disorder commonly characterized by chronic eosinophilic inflammation, remodeling and hyper responsiveness of the airways. However, the kinases like Phosphoinositide 3 kinase (PI3K) and Janus kinase 3 (JAK3) are involved in mast cell proliferation, activation, recruitment, migration, and prolonged survival of inflammatory cells. The present study was designed to evaluate the in-vivo comparative effects of two kinase inhibitors on airway inflammation and airway remodeling in acute and chronic models of asthma. Mice were sensitized twice intra-peritoneally and then challenged by inhalation with ovalbumin (OVA). They developed an extensive inflammatory response, goblet cell hyperplasia, collagen deposition, airway smooth muscle thickening similar to pathologies observed in human asthma. The effects of PI3K inhibitor (30 mg/kg, p.o), JAK3 inhibitor (30 mg/kg, p.o) and Dexamethasone (0.3 mg/kg) on airway inflammation and remodeling in OVA sensitized/challenged BALB/c mice were evaluated. Twenty-four hours after the final antigen challenge, bronchoalveolar lavage (BAL) and histological examinations were carried out. It was observed that kinase inhibitors significantly reduced airway inflammation as evidenced by the decrease in pro inflammatory cytokines in BALF and lung homogenate and inflammatory cell count in sensitized mice after allergen challenge. Lung histological analysis showed increased infiltration of inflammatory cells, hyperplasia of goblet cells and the collagen deposition, which were significantly reduced with kinase inhibitor. In conclusion, our data suggest that PI3K and JAK3 inhibitors showed promising alternative therapeutic activity in asthma, which might significantly counteract the airway inflammation in patients with allergic asthma.

  17. Role of phosphoinositide 3-kinase IA (PI3K-IA) activation in cardioprotection induced by ouabain preconditioning.

    PubMed

    Duan, Qiming; Madan, Namrata D; Wu, Jian; Kalisz, Jennifer; Doshi, Krunal Y; Haldar, Saptarsi M; Liu, Lijun; Pierre, Sandrine V

    2015-03-01

    Acute myocardial infarction, the clinical manifestation of ischemia-reperfusion (IR) injury, is a leading cause of death worldwide. Like ischemic preconditioning (IPC) induced by brief episodes of ischemia and reperfusion, ouabain preconditioning (OPC) mediated by Na/K-ATPase signaling protects the heart against IR injury. Class I PI3K activation is required for IPC, but its role in OPC has not been investigated. While PI3K-IB is critical to IPC, studies have suggested that ouabain signaling is PI3K-IA-specific. Hence, a pharmacological approach was used to test the hypothesis that OPC and IPC rely on distinct PI3K-I isoforms. In Langendorff-perfused mouse hearts, OPC was initiated by 4 min of ouabain 10 μM and IPC was triggered by 4 cycles of 5 min ischemia and reperfusion prior to 40 min of global ischemia and 30 min of reperfusion. Without affecting PI3K-IB, ouabain doubled PI3K-IA activity and Akt phosphorylation at Ser(473). IPC and OPC significantly preserved cardiac contractile function and tissue viability as evidenced by left ventricular developed pressure and end-diastolic pressure recovery, reduced lactate dehydrogenase release, and decreased infarct size. OPC protection was blunted by the PI3K-IA inhibitor PI-103, but not by the PI3K-IB inhibitor AS-604850. In contrast, IPC-mediated protection was not affected by PI-103 but was blocked by AS-604850, suggesting that PI3K-IA activation is required for OPC while PI3K-IB activation is needed for IPC. Mechanistically, PI3K-IA activity is required for ouabain-induced Akt activation but not PKCε translocation. However, in contrast to PKCε translocation which is critical to protection, Akt activity was not required for OPC. Further studies shall reveal the identity of the downstream targets of this new PI3K IA-dependent branch of OPC. These findings may be of clinical relevance in patients at risk for myocardial infarction with underlying diseases and/or medication that could differentially affect the

  18. Phosphatidylinositol (3,4) bisphosphate-specific phosphatases and effector proteins: A distinct branch of PI3K signaling.

    PubMed

    Li, Hongzhao; Marshall, Aaron J

    2015-09-01

    The ubiquitously expressed phosphoinositide 3-kinase (PI3K) family of lipid kinases control diverse cellular functions including cell survival, proliferation, metabolism and migration. Class I PI3Ks generate two distinct 3-phosphoinositide lipid messengers, PI(3,4,5)P3 (PIP3) and PI(3,4)P2, that recruit signaling effectors such as pleckstrin homology (PH) domain-containing proteins. Historically, the function of PI3K signaling has often been attributed to PIP3, with PI(3,4)P2 considered an inconsequential byproduct of PIP3 hydrolysis by SHIP phosphatases. However, accumulating evidence has demonstrated that PI(3,4)P2 directs a distinct branch of the PI3K pathway that regulates a variety of cellular processes with relevance to health and disease, such as B cell activation and autoantibody production, insulin sensitivity, neuronal dynamics, endocytosis and cell migration. Signaling through PI(3,4)P2 can be negatively regulated by inositol polyphosphate 4-phosphatases (INPP4A and INPP4B), which selectively degrade PI(3,4)P2. A number of signaling proteins that specifically bind to PI(3,4)P2 have been characterized, including the tandem PH domain-containing proteins (TAPP1 and TAPP2) and lamellipodin/RAPH1. A number of PIP3-binding proteins also bind to PI(3,4)P2, such as the protein kinase Akt/PKB, the most studied effector of PI3K signaling. Here, we review the current progress in understanding the functions and mechanisms of action of the PI(3,4)P2-specific phosphatases and binding proteins. A summary of available data addressing the relative contribution of PI(3,4)P2 versus PIP3 in regulation of Akt is provided to highlight the potential independent role of PI(3,4)P2 in regulating some PIP3-binding proteins. In summary, PI(3,4)P2-specific phosphatases and binding proteins are now firmly established players in cell biology, and this "neglected" phosphoinositide needs to take its place as one of the central components of the PI3K signaling pathway.

  19. Age-dependent accumulation of soluble amyloid beta (Abeta) oligomers reverses the neuroprotective effect of soluble amyloid precursor protein-alpha (sAPP(alpha)) by modulating phosphatidylinositol 3-kinase (PI3K)/Akt-GSK-3beta pathway in Alzheimer mouse model.

    PubMed

    Jimenez, Sebastian; Torres, Manuel; Vizuete, Marisa; Sanchez-Varo, Raquel; Sanchez-Mejias, Elisabeth; Trujillo-Estrada, Laura; Carmona-Cuenca, Irene; Caballero, Cristina; Ruano, Diego; Gutierrez, Antonia; Vitorica, Javier

    2011-05-27

    Neurotrophins, activating the PI3K/Akt signaling pathway, control neuronal survival and plasticity. Alterations in NGF, BDNF, IGF-1, or insulin signaling are implicated in the pathogenesis of Alzheimer disease. We have previously characterized a bigenic PS1×APP transgenic mouse displaying early hippocampal Aβ deposition (3 to 4 months) but late (17 to 18 months) neurodegeneration of pyramidal cells, paralleled to the accumulation of soluble Aβ oligomers. We hypothesized that PI3K/Akt/GSK-3β signaling pathway could be involved in this apparent age-dependent neuroprotective/neurodegenerative status. In fact, our data demonstrated that, as compared with age-matched nontransgenic controls, the Ser-9 phosphorylation of GSK-3β was increased in the 6-month PS1×APP hippocampus, whereas in aged PS1×APP animals (18 months), GSK-3β phosphorylation levels displayed a marked decrease. Using N2a and primary neuronal cell cultures, we demonstrated that soluble amyloid precursor protein-α (sAPPα), the predominant APP-derived fragment in young PS1×APP mice, acting through IGF-1 and/or insulin receptors, activated the PI3K/Akt pathway, phosphorylated the GSK-3β activity, and in consequence, exerted a neuroprotective action. On the contrary, several oligomeric Aβ forms, present in the soluble fractions of aged PS1×APP mice, inhibited the induced phosphorylation of Akt/GSK-3β and decreased the neuronal survival. Furthermore, synthetic Aβ oligomers blocked the effect mediated by different neurotrophins (NGF, BDNF, insulin, and IGF-1) and sAPPα, displaying high selectivity for NGF. In conclusion, the age-dependent appearance of APP-derived soluble factors modulated the PI3K/Akt/GSK-3β signaling pathway through the major neurotrophin receptors. sAPPα stimulated and Aβ oligomers blocked the prosurvival signaling. Our data might provide insights into the selective vulnerability of specific neuronal groups in Alzheimer disease.

  20. Targeting the Phosphatidylinositol-3-kinase Pathway in Gastric Cancer: Can Omics Improve Outcomes?

    PubMed Central

    Tran, Phu; Nguyen, Cham

    2016-01-01

    Phosphatidylinositol-3-kinase (PI3K) pathway signaling is an established oncogenic signal transduction pathway implicated in multiple malignancies. Therapeutic targeting of PI3K pathway components has improved outcomes in chronic lymphocytic leukemia, kidney cancer, breast cancer, and neuroendocrine tumors. Gastric cancers harbor some of the highest rates of oncogenic alterations in PI3K but attempts to translate this genomic observation have met with limited clinical success and novel approaches are needed. In the following review we discuss PI3K signaling, previous preclinical and clinical investigations in gastric cancer, and discuss future strategies aimed at overcoming resistance and improving efficacy. Identification and refinement of molecular tumor subtypes, development of predictive biomarkers along, and rational drug combination strategies are key to capitalizing on the therapeutic potential of PI3K pathway directed therapies in gastric cancers. PMID:27915478

  1. Malignant Pleural Effusion and ascites Induce Epithelial-Mesenchymal Transition and Cancer Stem-like Cell Properties via the Vascular Endothelial Growth Factor (VEGF)/Phosphatidylinositol 3-Kinase (PI3K)/Akt/Mechanistic Target of Rapamycin (mTOR) Pathway.

    PubMed

    Yin, Tao; Wang, Guoping; He, Sisi; Shen, Guobo; Su, Chao; Zhang, Yan; Wei, Xiawei; Ye, Tinghong; Li, Ling; Yang, Shengyong; Li, Dan; Guo, Fuchun; Mo, Zeming; Wan, Yang; Ai, Ping; Zhou, Xiaojuan; Liu, Yantong; Wang, Yongsheng; Wei, Yuquan

    2016-12-23

    Malignant pleural effusion (PE) and ascites, common clinical manifestations in advanced cancer patients, are associated with a poor prognosis. However, the biological characteristics of malignant PE and ascites are not clarified. Here we report that malignant PE and ascites can induce a frequent epithelial-mesenchymal transition program and endow tumor cells with stem cell properties with high efficiency, which promotes tumor growth, chemoresistance, and immune evasion. We determine that this epithelial-mesenchymal transition process is mainly dependent on VEGF, one initiator of the PI3K/Akt/mechanistic target of rapamycin (mTOR) pathway. From the clinical observation, we define a therapeutic option with VEGF antibody for malignant PE and ascites. Taken together, our findings clarify a novel biological characteristic of malignant PE and ascites in cancer progression and provide a promising and available strategy for cancer patients with recurrent/refractory malignant PE and ascites. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Cudraflavone C Induces Tumor-Specific Apoptosis in Colorectal Cancer Cells through Inhibition of the Phosphoinositide 3-Kinase (PI3K)-AKT Pathway

    PubMed Central

    Soo, Hsien-Chuen; Chung, Felicia Fei-Lei; Lim, Kuan-Hon; Yap, Veronica Alicia; Bradshaw, Tracey D.; Hii, Ling-Wei; Tan, Si-Hoey; See, Sze-Jia; Tan, Yuen-Fen; Leong, Chee-Onn

    2017-01-01

    Cudraflavone C (Cud C) is a naturally-occurring flavonol with reported anti-proliferative activities. However, the mechanisms by which Cud C induced cytotoxicity have yet to be fully elucidated. Here, we investigated the effects of Cud C on cell proliferation, caspase activation andapoptosis induction in colorectal cancer cells (CRC). We show that Cud C inhibits cell proliferation in KM12, Caco-2, HT29, HCC2998, HCT116 and SW48 CRC but not in the non-transformed colorectal epithelial cells, CCD CoN 841. Cud C induces tumor-selective apoptosis via mitochondrial depolarization and activation of the intrinsic caspase pathway. Gene expression profiling by microarray analyses revealed that tumor suppressor genes EGR1, HUWE1 and SMG1 were significantly up-regulated while oncogenes such as MYB1, CCNB1 and GPX2 were down-regulated following treatment with Cud C. Further analyses using Connectivity Map revealed that Cud C induced a gene signature highly similar to that of protein synthesis inhibitors and phosphoinositide 3-kinase (PI3K)-AKT inhibitors, suggesting that Cud C might inhibit PI3K-AKT signaling. A luminescent cell free PI3K lipid kinase assay revealed that Cud C significantly inhibited p110β/p85α PI3K activity, followed by p120γ, p110δ/p85α, and p110α/p85α PI3K activities. The inhibition by Cud C on p110β/p85α PI3K activity was comparable to LY-294002, a known PI3K inhibitor. Cud C also inhibited phosphorylation of AKT independent of NFκB activity in CRC cells, while ectopic expression of myristoylated AKT completely abrogated the anti-proliferative effects, and apoptosis induced by Cud C in CRC. These findings demonstrate that Cud C induces tumor-selective cytotoxicity by targeting the PI3K-AKT pathway. These findings provide novel insights into the mechanism of action of Cud C, and indicate that Cud C further development of Cud C derivatives as potential therapeutic agents is warranted. PMID:28107519

  3. ZSTK474, a novel phosphatidylinositol 3-kinase inhibitor identified using the JFCR39 drug discovery system.

    PubMed

    Kong, De-xin; Yamori, Takao

    2010-09-01

    JFCR39 is an informatic anticancer drug discovery system that utilizes a panel of 39 human cancer cells coupled with a drug-activity database. This system not only provides disease-oriented information but can also predict the mechanism of action of a given antitumor agent. Development of a phosphatidylinositol 3-kinase (PI3K) inhibitor as an anticancer drug candidate has attracted a great deal of attention from both academia and industry because PI3K is known to be closely involved in carcinogenesis. ZSTK474 was identified as a PI3K inhibitor using JFCR39 system in combination with COMPARE analysis program. These findings were based on the similar fingerprint (growth inhibition profiles for JFCR39 human cancer cell line panel) with that of a classical PI3K inhibitor LY294002. Biochemical experiments confirmed ZSTK474 to be a potent pan-class I PI3K inhibitor, with high selectivity over other classes of PI3K and protein kinases. We previously reported the in vitro and in vivo antitumor efficacy of ZSTK474, together with the G(0)/G(1) arrest and antiangiogenic activity. Here, we review the JFCR39 system and summarize recent studies on PI3K biology and the development of PI3K inhibitors before discussing ZSTK474 in some detail.

  4. Phosphatidylinositol 3-kinase activates ERK in primary sensory neurons and mediates inflammatory heat hyperalgesia through TRPV1 sensitization.

    PubMed

    Zhuang, Zhi-Ye; Xu, Haoxing; Clapham, David E; Ji, Ru-Rong

    2004-09-22

    Although the PI3K (phosphatidylinositol 3-kinase) pathway typically regulates cell growth and survival, increasing evidence indicates the involvement of this pathway in neural plasticity. It is unknown whether the PI3K pathway can mediate pain hypersensitivity. Intradermal injection of capsaicin and NGF produce heat hyperalgesia by activating their respective TRPV1 (transient receptor potential vanilloid receptor-1) and TrkA receptors on nociceptor sensory nerve terminals. We examined the activation of PI3K in primary sensory DRG neurons by these inflammatory agents and the contribution of PI3K activation to inflammatory pain. We further investigated the correlation between the PI3K and the ERK (extracellular signal-regulated protein kinase) pathway. Capsaicin and NGF induce phosphorylation of the PI3K downstream target AKT (protein kinase B), which is blocked by the PI3K inhibitors LY294002 and wortmannin, indicative of the activation of PI3K by both agents. ERK activation by capsaicin and NGF was also blocked by PI3K inhibitors. Similarly, intradermal capsaicin in rats activated PI3K and ERK in C-fiber DRG neurons and epidermal nerve fibers. Injection of PI3K or MEK (ERK kinase) inhibitors into the hindpaw attenuated capsaicin- and NGF-evoked heat hyperalgesia but did not change basal heat sensitivity. Furthermore, PI3K, but not ERK, inhibition blocked early induction of hyperalgesia. In acutely dissociated DRG neurons, the capsaicin-induced TRPV1 current was strikingly potentiated by NGF, and this potentiation was completely blocked by PI3K inhibitors and primarily suppressed by MEK inhibitors. Therefore, PI3K induces heat hyperalgesia, possibly by regulating TRPV1 activity, in an ERK-dependent manner. The PI3K pathway also appears to play a role that is distinct from ERK by regulating the early onset of inflammatory pain.

  5. Control of neurite outgrowth and growth cone motility by phosphatidylinositol-3-kinase.

    PubMed

    Tornieri, Karine; Welshhans, Kristy; Geddis, Matthew S; Rehder, Vincent

    2006-04-01

    Phosphatidylinositol-3-kinase (PI-3K) has been reported to affect neurite outgrowth both in vivo and in vitro. Here we investigated the signaling pathways by which PI-3K affects neurite outgrowth and growth cone motility in identified snail neurons in vitro. Inhibition of PI-3K with wortmannin (2 microM) or LY 294002 (25 microM) resulted in a significant elongation of filopodia and in a slow-down of neurite outgrowth. Experiments using cytochalasin and blebbistatin, drugs that interfere with actin polymerization and myosin II activity, respectively, demonstrated that filopodial elongation resulting from PI-3K inhibition was dependent on actin polymerization. Inhibition of strategic kinases located downstream of PI-3K, such as Akt, ROCK, and MEK, also caused significant filopodial elongation and a slow-down in neurite outgrowth. Another growth cone parameter, filopodial number, was not affected by inhibition of PI-3K, Akt, ROCK, or MEK. A detailed study of growth cone behavior showed that the filopodial elongation induced by inhibiting PI-3K, Akt, ROCK, and MEK was achieved by increasing two motility parameters: the rate with which filopodia extend (extension rate) and the time that filopodia spend elongating. Whereas the inhibition of ROCK or Akt (both activated by the lipid kinase activity of PI-3K) and MEK (activated by the protein kinase activity of PI-3K) had additive effects, simultaneous inhibition of Akt and ROCK showed no additive effect. We further demonstrate that the effects on filopodial dynamics investigated were calcium-independent. Taken together, our results suggest that inhibition of PI-3K signaling results in filopodial elongation and a slow-down of neurite advance, reminiscent of growth cone searching behavior.

  6. Selective Sparing of Human Tregs by Pharmacologic Inhibitors of the Phosphatidylinositol 3-Kinase and MEK Pathways

    PubMed Central

    Zwang, N. A.; Zhang, R.; Germana, S.; Fan, M. Y.; Hastings, W. D.; Cao, A.; Turka, L. A.

    2016-01-01

    Phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase/extracellular signal-regulated (MEK) signaling are central to the survival and proliferation of many cell types. Multiple lines of investigation in murine models have shown that control of the PI3K pathway is particularly important for regulatory T cell (Treg) stability and function. PI3K and MEK inhibitors are being introduced into the clinic, and we hypothesized that pharmacologic inhibition of PI3K, and possibly MEK, in mixed cultures of human mononuclear cells would preferentially affect CD4+ and CD8+ lymphocytes compared with Tregs. We tested this hypothesis using four readouts: proliferation, activation, functional suppression, and signaling. Results showed that Tregs were less susceptible to inhibition by both δ and α isoform–specific PI3K inhibitors and by an MEK inhibitor compared with their conventional CD4+ and CD8+ counterparts. These studies suggest less functional reliance on PI3K and MEK signaling in Tregs compared with conventional CD4+ and CD8+ lymphocytes. Therefore, the PI3K and MEK pathways are attractive pharmacologic targets for transplantation and treatment of autoimmunity. PMID:27017850

  7. Advances in the researches on the biological activities and inhibitors of phosphatidylinositol 3-kinase.

    PubMed

    Tang, Jian-Feng; Wen, Qing; Sun, Jian; Zhang, Wei-Ming; Zhu, Hai-Liang

    2014-06-01

    The PI3K/AKT/mTOR pathway is an intracellular signaling pathway, being important in apoptosis hence cancer such as breast cancer and non-small-cell lung cancer. It signaling axis controls cell proliferation and survival and has achieved major importance as a target for cancer therapy. The serine/threonine kinase Akt (also known as protein kinase B or PKB), since its initial discovery as a protooncogene, has become a major focus of attention because of its critical regulatory role in diverse cellular processes, including cancer progression and insulin metabolism. The Akt cascade is activated by receptor tyrosine kinases, integrins, B and T cell receptors, cytokine receptors, G protein coupled receptors and other stimuli that induce the production of phosphatidylinositol 3,4,5 triphosphates (PtdIns(3,4,5)P3) by phosphoinositide 3-kinase (PI3K). Therefore, PI3K plays an important role in in numerous cellular functions such as cell growth, proliferation, differentiation, motility, survival and intracellular trafficking. In this review, we introduced the structure of the PI3K, and then focused on its biological activities. In addition, we reviewed the advances in the researches of PI3K as well as related inhibitors over the last couple of decades. Finally, we also discussed the prospect and developmental trend of phosphatidylinositol 3-kinase as antitumor agents.

  8. Fenofibrate pre-treatment suppressed inflammation by activating phosphoinositide 3 kinase/protein kinase B (PI3K/Akt) signaling in renal ischemia-reperfusion injury.

    PubMed

    Yang, Feng-jie; He, Yong-hua; Zhou, Jian-hua

    2015-02-01

    The aim of this study was to investigate the possible beneficial effects of Fenofibrate on renal ischemia-reperfusion injury (IRI) in mice and its potential mechanism. IRI was induced by bilateral renal ischemia for 60 min followed by reperfusion for 24 h. Eighteen male C57BL/6 mice were randomly divided into three groups: sham-operated group (sham), IRI+saline group (IRI group), IRI+Fenofibrate (FEN) group. Normal saline or Fenofibrate (3 mg/kg) was intravenously injected 60 min before renal ischemia in IRI group and FEN group, respectively. Blood samples and renal tissues were collected at the end of reperfusion. The renal function, histopathologic changes, and the expression levels of pro-inflammatory cytokines [interleukin-8 (IL-8), tumor necrosis factor alpha (TNF-α) and IL-6] in serum and renal tissue homogenate were assessed. Moreover, the effects of Fenofibrate on activating phosphoinositide 3 kinase/protein kinase B (PI3K/Akt) signaling and peroxisome proliferator-activated receptor-α (PPAR-α) were also measured in renal IRI. The results showed that plasma levels of blood urea nitrogen and creatinine, histopathologic scores and the expression levels of TNF-α, IL-8 and IL-6 were significantly lower in FEN group than in IRI group. Moreover, Fenofibrate pretreatment could further induce PI3K/Akt signal pathway and PPAR-α activation following renal IRI. These findings indicated PPAR-α activation by Fenofibrate exerts protective effects on renal IRI in mice by suppressing inflammation via PI3K/Akt activation. Thus, Fenofibrate could be a novel therapeutic alternative in renal IRI.

  9. Progress in the Preclinical Discovery and Clinical Development of Class I and Dual Class I/IV Phosphoinositide 3-Kinase (PI3K) Inhibitors

    PubMed Central

    Shuttleworth, S.J; Silva, F.A; Cecil, A.R.L; Tomassi, C.D; Hill, T.J; Raynaud, F.I; Clarke, P.A; Workman, P

    2011-01-01

    The phosphoinositide 3-kinases (PI3Ks) constitute an important family of lipid kinase enzymes that control a range of cellular processes through their regulation of a network of signal transduction pathways, and have emerged as important therapeutic targets in the context of cancer, inflammation and cardiovascular diseases. Since the mid-late 1990s, considerable progress has been made in the discovery and development of small molecule ATP-competitive PI3K inhibitors, a number of which have entered early phase human trials over recent years from which key clinical results are now being disclosed. This review summarizes progress made to date, primarily on the discovery and characterization of class I and dual class I/IV subtype inhibitors, together with advances that have been made in translational and clinical research, notably in cancer. PMID:21649578

  10. Sulforaphane prevents human platelet aggregation through inhibiting the phosphatidylinositol 3-kinase/Akt pathway.

    PubMed

    Chuang, Wen-Ying; Kung, Po-Hsiung; Kuo, Chih-Yun; Wu, Chin-Chung

    2013-06-01

    Sulforaphane, a dietary isothiocyanate found in cruciferous vegetables, has been shown to exert beneficial effects in animal models of cardiovascular diseases. However, its effect on platelet aggregation, which is a critical factor in arterial thrombosis, is still unclear. In the present study, we show that sulforaphane inhibited human platelet aggregation caused by different receptor agonists, including collagen, U46619 (a thromboxane A2 mimic), protease-activated receptor 1 agonist peptide (PAR1-AP), and an ADP P2Y12 receptor agonist. Moreover, sulforaphane significantly reduced thrombus formation on a collagen-coated surface under whole blood flow conditions. In exploring the underlying mechanism, we found that sulforaphane specifically prevented phosphatidylinositol 3-kinase (PI3K)/Akt signalling, without markedly affecting other signlaling pathways involved in platelet aggregation, such as protein kinase C activation, calcium mobilisation, and protein tyrosine phosphorylation. Although sulforaphane did not directly inhibit the catalytic activity of PI3K, it caused ubiquitination of the regulatory p85 subunit of PI3K, and prevented PI3K translocation to membranes. In addition, sulforaphane caused ubiquitination and degradation of phosphoinositide-dependent kinase 1 (PDK1), which is required for Akt activation. Therefore, sulforaphane is able to inhibit the PI3K/Akt pathway at two distinct sites. In conclusion, we have demonstrated that sulforaphane prevented platelet aggregation and reduced thrombus formation in flow conditions; our data also support that the inhibition of the PI3K/Akt pathway by sulforaphane contributes it antiplatelet effects.

  11. New drug development in head and neck squamous cell carcinoma: The PI3-K inhibitors.

    PubMed

    De Felice, Francesca; Guerrero Urbano, Teresa

    2017-04-01

    Over the last few years a number of new different compounds have been developed. They include phosphatidylinositol 3-kinase (PI3-K) inhibitors. Deregulation within the PI3-K pathway is common in head neck squamous cell carcinoma (HNSCC) and it represents a growing area of research. PI3-K inhibitors, including BKM120, PX-866 and BYL719, are being tested in several phase I and phase II studies in patients with locally advanced, recurrent or metastatic disease. This review provides an update of published clinical trials and highlights the challenges of PI3-K inhibitors in HNSCC.

  12. Phosphatidylinositol 3-kinase inhibition broadly sensitizes glioblastoma cells to death receptor- and drug-induced apoptosis.

    PubMed

    Opel, Daniela; Westhoff, Mike-Andrew; Bender, Ariane; Braun, Veit; Debatin, Klaus-Michael; Fulda, Simone

    2008-08-01

    The aberrant activity of the phosphatidylinositol 3-kinase (PI3K) pathway has been reported to correlate with adverse clinical outcome in human glioblastoma in vivo. However, the question of how this survival network can be successfully targeted to restore the sensitivity of glioblastoma to apoptosis induction has not yet been answered. Here, we report that inhibition of PI3K by LY294002 broadly sensitizes wild-type and mutant PTEN glioblastoma cells to both death receptor- and chemotherapy-induced apoptosis, whereas mammalian target of rapamycin (mTOR) inhibition is not sufficient to restore apoptosis sensitivity. LY294002 significantly enhances apoptosis triggered by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), agonistic anti-CD95 antibodies, or several anticancer drugs (i.e., doxorubicin, etoposide, and vincristine) in a highly synergistic manner. In addition, LY294002 cooperates with TRAIL or doxorubicin to suppress colony formation, thus also showing a strong effect on long-term survival. Similarly, genetic knockdown of PI3K subunits p110alpha and/or p110beta by RNA interference (RNAi) primes glioblastoma cells for TRAIL- or doxorubicin-mediated apoptosis. In contrast to PI3K inhibition, pharmacologic or genetic blockade of mTOR by RAD001 (everolimus), rapamycin, or RNAi fails to enhance TRAIL- or doxorubicin-induced apoptosis. Analysis of apoptosis pathways reveals that PI3K inhibition acts in concert with TRAIL or doxorubicin to trigger mitochondrial membrane permeabilization, caspase activation, and caspase-dependent apoptosis, which are abolished by the caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone. Most importantly, PI3K inhibition by LY294002 sensitizes primary cultured glioblastoma cells obtained from surgical specimens to TRAIL- or chemotherapy-induced cell death. By showing that PI3K inhibition broadly primes glioblastoma cells for apoptosis, our findings provide the rationale for using PI3K inhibitors in

  13. Phosphatidylinositol 3-kinase pathway regulates sperm viability but not capacitation on boar spermatozoa.

    PubMed

    Aparicio, I M; Bragado, M J; Gil, M C; Garcia-Herreros, M; Gonzalez-Fernandez, L; Tapia, J A; Garcia-Marin, L J

    2007-08-01

    Phosphatidylinositol 3-kinase (PI3-K) plays an important role in cell survival in somatic cells and recent data pointed out a role for this kinase in sperm capacitation and acrosome reaction (AR). This study was undertaken to evaluate the role of PI3-K pathway on porcine spermatozoa capacitation, AR, and viability using two unrelated PI3-K inhibitors, LY294002 and wortmannin. In boar spermatozoa, we have identified the presence of PDK1, PKB/Akt, and PTEN, three of the main key components of the PI3-K pathway. Incubation of boar sperm in a capacitating medium (TCM) caused a significant increase in the percentage of capacitated (25 +/- 2 to 34 +/- 1% P < 0.05, n = 6) and acrosome reacted (1 +/- 1 to 11 +/- 1% P < 0.01, n = 6) spermatozoa compared with sperm in basal medium (TBM). Inhibition of PI3-K did affect neither the capacitation status nor AR nor protein p32 tyrosine phosphorylation of boar spermatozoa incubated in TBM or TCM. Boar sperm viability in TBM was significantly decreased by 40 and 20% after pretreatment with LY294002 or wortmannin, respectively. Similar results were observed after incubation of boar spermatozoa in TCM. Treatment of boar spermatozoa with the analog of cAMP, 8Br-cAMP significantly prevented the reduction on sperm viability. Our results provide evidence for an important role of the PI3-K pathway in the regulation of boar sperm viability and suggests that other signaling pathways different from PI3-K must be activated downstream of cAMP to contribute to regulation of sperm viability. Finally, in our conditions the PI3-K pathway seems not related with boar sperm capacitation or AR.

  14. A genomewide overexpression screen identifies genes involved in the phosphatidylinositol 3-kinase pathway in the human protozoan parasite Entamoeba histolytica.

    PubMed

    Koushik, Amrita B; Welter, Brenda H; Rock, Michelle L; Temesvari, Lesly A

    2014-03-01

    Entamoeba histolytica is a protozoan parasite that causes amoebic dysentery and liver abscess. E. histolytica relies on motility, phagocytosis, host cell adhesion, and proteolysis of extracellular matrix for virulence. In eukaryotic cells, these processes are mediated in part by phosphatidylinositol 3-kinase (PI3K) signaling. Thus, PI3K may be critical for virulence. We utilized a functional genomics approach to identify genes whose products may operate in the PI3K pathway in E. histolytica. We treated a population of trophozoites that were overexpressing genes from a cDNA library with a near-lethal dose of the PI3K inhibitor wortmannin. This screen was based on the rationale that survivors would be overexpressing gene products that directly or indirectly function in the PI3K pathway. We sequenced the overexpressed genes in survivors and identified a cDNA encoding a Rap GTPase, a protein previously shown to participate in the PI3K pathway. This supports the validity of our approach. Genes encoding a coactosin-like protein, EhCoactosin, and a serine-rich E. histolytica protein (SREHP) were also identified. Cells overexpressing EhCoactosin or SREHP were also less sensitive to a second PI3K inhibitor, LY294002. This corroborates the link between these proteins and PI3K. Finally, a mutant cell line with an increased level of phosphatidylinositol (3,4,5)-triphosphate, the product of PI3K activity, exhibited increased expression of SREHP and EhCoactosin. This further supports the functional connection between these proteins and PI3K in E. histolytica. To our knowledge, this is the first forward-genetics screen adapted to reveal genes participating in a signal transduction pathway in this pathogen.

  15. Regulation of the PI3K pathway through a p85α monomer–homodimer equilibrium | Office of Cancer Genomics

    Cancer.gov

    The canonical action of the p85α regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is to associate with the p110α catalytic subunit to allow stimuli-dependent activation of the PI3K pathway. We elucidate a p110α-independent role of homodimerized p85α in the positive regulation of PTEN stability and activity.

  16. Predicting the structures of complexes between phosphoinositide 3-kinase (PI3K) and romidepsin-related compounds for the drug design of PI3K/histone deacetylase dual inhibitors using computational docking and the ligand-based drug design approach.

    PubMed

    Oda, Akifumi; Saijo, Ken; Ishioka, Chikashi; Narita, Koichi; Katoh, Tadashi; Watanabe, Yurie; Fukuyoshi, Shuichi; Takahashi, Ohgi

    2014-11-01

    Predictions of the three-dimensional (3D) structures of the complexes between phosphoinositide 3-kinase (PI3K) and two inhibitors were conducted using computational docking and the ligand-based drug design approach. The obtained structures were refined by structural optimizations and molecular dynamics (MD) simulations. The ligands were located deep inside the ligand binding pocket of the p110α subunit of PI3K, and the hydrogen bond formations and hydrophobic effects of the surrounding amino acids were predicted. Although rough structures were obtained for the PI3K-inhibitor complexes before the MD simulations, the refinement of the structures by these simulations clarified the hydrogen bonding patterns of the complexes.

  17. Phosphatidylinositol-3 kinase-dependent translational regulation of Id1 involves the PPM1G phosphatase

    PubMed Central

    Xu, Kaiming; Wang, Lanfang; Feng, Wei; Feng, Yue; Shu, Hui-Kuo G.

    2016-01-01

    Id1 is a helix-loop-helix transcriptional modulator that increases the aggressiveness of malignant glial neoplasms. Since most glioblastomas (GBMs) show increased phosphatidylinositol-3 kinase (PI-3K) signaling, we sought to determine whether this pathway regulates Id1 expression. Higher basal Id1 expression correlates with dysregulated PI-3K signaling in multiple established GBM cell lines. Further characterization of PI-3K-dependent Id1 regulation reveals that chemical or genetic inhibition of PI-3K signaling reduces Id1 protein but not mRNA expression. Overall, PI-3K signaling appears to enhance Id1 translation with no significant effect on its stability. PI-3K signaling is known to regulate protein translation through mTORC1-dependent phosphorylation of 4E-BP1, which reduces its association with and inhibition of the translation initiation factor eIF4E. Interestingly, while inhibition of PI-3K and AKT lowers 4E-BP1 phosphorylation and expression of Id1 in all cases, inhibition of TORC1 with rapamycin does not consistently have a similar effect suggesting an alternative mechanism for PI-3K-dependent regulation of Id1 translation. We now identify a potential role for the serine-threonine phosphatase PPM1G in translational regulation of Id1 protein expression. PPM1G knockdown by siRNA increase both 4E-BP1 phosphorylation and Id1 expression and PPM1G and 4E-BP1 co-associates in GBM cells. Furthermore, PPM1G is a phosphoprotein and this phosphorylation appears to be regulated by PI-3K activity. Finally, PI-3K inhibition increases PPM1G activity when assessed by an in vitro phosphatase assay. Our findings provide the first evidence that the PI-3K/AKT signaling pathway modulates PPM1G activity resulting in a shift in the balance between hyper- and hypo-phosphorylated 4E-BP1 and translational regulation of Id1 expression. PMID:27065332

  18. Phosphatidylinositol 3-kinase mediates the ability of retinol to decrease colorectal cancer cell invasion.

    PubMed

    Lengyel, Jennifer N Griffin; Park, Eun Young; Brunson, Anna R; Pinali, Daniel; Lane, Michelle A

    2014-01-01

    Previously, we showed that retinol (vitamin A) decreased both colorectal cancer cell invasion and phosphatidylinositol 3-kinase (PI3K) activity through a retinoic acid receptor-independent mechanism. Here, we determined if these phenomena were related by using parental HCT-116 cells that harbor 1 allele of wild-type PI3K and 1 allele of constitutively active (ca) PI3K and 2 mutant HCT-116 cell lines homozygous for caPI3K. In vitro, treatment of parental HCT-116 cells with 10 μM retinol reduced cell invasion whereas treatment of mutant HCT-116 cell lines with retinol did not. Treatment with 10 μM retinol also decreased the activity of matrixmetalloproteinase-9 and increased tissue inhibitor of matrixmetalloproteinase-I levels in parental, but not mutant, HCT-116 cells. Finally, parental or mutant cells were intrasplenically injected into athymic mice consuming diets with or without supplemental vitamin A. As expected, vitamin A supplementation tended (P = 0.18) to reduce the incidence of metastases in mice injected with the parental cell line and consuming the supplemented diet. In contrast, metastatic incidence was not affected (P = 1.00) by vitamin A supplementation in mice injected with mutant cells. These data indicate that the capacity of retinol to inhibit PI3K activity confers its ability to decrease colorectal cancer metastasis.

  19. Eupafolin suppresses prostate cancer by targeting phosphatidylinositol 3-kinase-mediated Akt signaling.

    PubMed

    Liu, Kangdong; Park, Chanmi; Chen, Hanyong; Hwang, Joonsung; Thimmegowda, N R; Bae, Eun Young; Lee, Ki Won; Kim, Hong-Gyum; Liu, Haidan; Soung, Nak Kyun; Peng, Cong; Jang, Jae Hyuk; Kim, Kyoon Eon; Ahn, Jong Seog; Bode, Ann M; Dong, Ziming; Kim, Bo Yeon; Dong, Zigang

    2015-09-01

    Phosphatase and tensin homolog (PTEN) loss or mutation consistently activates the phosphatidylinositol 3-kinase (PI3-K)/Akt signaling pathway, which contributes to the progression and invasiveness of prostate cancer. Furthermore, the PTEN/PI3-K/Akt and Ras/MAPK pathways cooperate to promote the epithelial-mesenchymal transition (EMT) and metastasis initiated from prostate stem/progenitor cells. For these reasons, the PTEN/PI3-K/Akt pathway is considered as an attractive target for both chemoprevention and chemotherapy. Herein we report that eupafolin, a natural compound found in common sage, inhibited proliferation of prostate cancer cells. Protein content analysis indicated that phosphorylation of Akt and its downstream kinases was inhibited by eupafolin treatment. Pull-down assay and in vitro kinase assay results indicated that eupafolin could bind with PI3-K and attenuate its kinase activity. Eupafolin also exhibited tumor suppressive effects in vivo in an athymic nude mouse model. Overall, these results suggested that eupafolin exerts antitumor effects by targeting PI3-K. © 2014 Wiley Periodicals, Inc.

  20. Eupafolin suppresses prostate cancer by targeting phosphatidylinositol 3-kinase-mediated Akt signaling

    PubMed Central

    Liu, Kangdong; Park, Chanmi; Chen, Hanyong; Hwang, Joonsung; Thimmegowda, N.R.; Bae, Eun Young; Lee, Ki Won; Kim, Hong-Gyum; Liu, Haidan; Soung, Nak Kyun; Peng, Cong; Jang, Jae Hyuk; Kim, Kyoon Eon; Ahn, Jong Seog; Bode, Ann M.; Dong, Ziming; Kim, Bo Yeon; Dong, Zigang

    2014-01-01

    Phosphatase and tensin homolog (PTEN) loss or mutation consistently activates the phosphatidylinositol 3-kinase (PI3-K)/Akt signaling pathway, which contributes to the progression and invasiveness of prostate cancer. Furthermore, the PTEN/PI3-K/Akt and Ras/MAPK pathways cooperate to promote the epithelial-mesenchymal transition (EMT) and metastasis initiated from prostate stem/progenitor cells. For these reasons, the PTEN/PI3-K/Akt pathway is considered as an attractive target for both chemoprevention and chemotherapy. Herein we report that eupafolin, a natural compound found in common sage, inhibited proliferation of prostate cancer cells. Protein content analysis indicated that phosphorylation of Akt and its downstream kinases was inhibited by eupafolin treatment. Pull-down assay and in vitro kinase assay results indicated that eupafolin could bind with PI3-K and attenuate its kinase activity. Eupafolin also exhibited tumor suppressive effects in vivo in an athymic nude mouse model. Overall, these results suggested that eupafolin exerts antitumor effects by targeting PI3-K. PMID:24700667

  1. Phosphatidylinositol 3-kinase inhibition potentiates glucocorticoid response in B-cell acute lymphoblastic leukemia.

    PubMed

    Evangelisti, Cecilia; Cappellini, Alessandra; Oliveira, Mariana; Fragoso, Rita; Barata, João T; Bertaina, Alice; Locatelli, Franco; Simioni, Carolina; Neri, Luca M; Chiarini, Francesca; Lonetti, Annalisa; Buontempo, Francesca; Orsini, Ester; Pession, Andrea; Manzoli, Lucia; Martelli, Alberto Maria; Evangelisti, Camilla

    2017-08-04

    Despite remarkable progress in polychemotherapy protocols, pediatric B-cell acute lymphoblastic leukemia (B-ALL) remains fatal in around 20% of cases. Hence, novel targeted therapies are needed for patients with poor prognosis. Glucocorticoids (GCs) are drugs commonly administrated for B-ALL treatment. Activation of the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin signaling pathway is frequently observed in B-ALL and contributes to GC-resistance. Here, we analyzed for the first time to our knowledge, the therapeutic potential of pan and isoform-selective PI3K p110 inhibitors, alone or combined with dexamethasone (DEX), in B-ALL leukemia cell lines and patient samples. We found that a pan PI3K p110 inhibitor displayed the most powerful cytotoxic effects in B-ALL cells, by inducing cell cycle arrest and apoptosis. Both a pan PI3K p110 inhibitor and a dual γ/δ PI3K p110 inhibitor sensitized B-ALL cells to DEX by restoring nuclear translocation of the GC receptor and counteracted stroma-induced DEX-resistance. Finally, gene expression analysis documented that, on one hand the combination consisting of a pan PI3K p110 inhibitor and DEX strengthened the DEX-induced up- or down-regulation of several genes involved in apoptosis, while on the other, it rescued the effects of genes that might be involved in GC-resistance. Overall, our findings strongly suggest that PI3K p110 inhibition could be a promising strategy for treating B-ALL patients by improving GC therapeutic effects and/or overcoming GC-resistance. © 2017 Wiley Periodicals, Inc.

  2. H1047R phosphatidylinositol 3-kinase mutant enhances HER2-mediated transformation via heregulin production and activation of HER3

    PubMed Central

    Chakrabarty, Anindita; Rexer, Brent N.; Wang, Shizhen Emily; Cook, Rebecca S.; Engelman, Jeffrey A.; Arteaga, Carlos, L.

    2010-01-01

    Hyperactivation of phosphatidylinositol-3 kinase (PI3K) can occur as a result of somatic mutations in PIK3CA, the gene encoding the p110α subunit of PI3K. The HER2 oncogene is amplified in 25% of all breast cancers and some of these tumors also harbor PIK3CA mutations. We examined mechanisms by which mutant PI3K can enhance transformation and confer resistance to HER2-directed therapies. We introduced the PI3K mutations E545K and H1047R in MCF10A human mammary epithelial cells that also overexpress HER2. Both mutants conferred a gain of function to MCF10A/HER2 cells. Expression of H1047R PI3K but not E545K PI3K markedly upregulated the HER3/HER4 ligand heregulin (HRG). HRG siRNA inhibited growth of H1047R but not E545K-expressing cells and synergized with the HER2 inhibitors trastuzumab and lapatinib. The PI3K inhibitor BEZ235 markedly inhibited HRG and pAKT levels and, in combination with lapatinib, completely inhibited growth of cells expressing H1047R PI3K. These observations suggest that PI3K mutants enhance HER2-mediated transformation by amplifying the ligand-induced signaling output of the ErbB network. This also counteracts the full effect of therapeutic inhibitors of HER2. These data also suggest that mammary tumors that contain both HER2 gene amplification and PIK3CA mutations should be treated with a combination of HER2 and PI3K inhibitors. PMID:20581867

  3. Pharmacodynamic Biomarker Development for PI3K Pathway Therapeutics

    PubMed Central

    Josephs, Debra H.; Sarker, Debashis

    2015-01-01

    The phosphatidylinositol 3-kinase (PI3K) signaling pathway is integral to many essential cell processes, including cell growth, differentiation, proliferation, motility, and metabolism. Somatic mutations and genetic amplifications that result in activation of the pathway are frequently detected in cancer. This has led to the development of rationally designed therapeutics targeting key members of the pathway. Critical to the successful development of these drugs are pharmacodynamic biomarkers that aim to define the degree of target and pathway inhibition. In this review, we discuss the pharmacodynamic biomarkers that have been utilized in early-phase clinical trials of PI3K pathway inhibitors. We focus on the challenges related to development and interpretation of these assays, their optimal integration with pharmacokinetic and predictive biomarkers, and future strategies to ensure successful development of PI3K pathway inhibitors within a personalized medicine paradigm for cancer. PMID:26917948

  4. Structure-Based Design of a Novel Series of Potent, Selective Inhibitors of the Class I Phosphatidylinositol 3-Kinases

    SciTech Connect

    Smith, Adrian L.; D’Angelo, Noel D.; Bo, Yunxin Y.; Booker, Shon K.; Cee, Victor J.; Herberich, Brad; Hong, Fang-Tsao; Jackson, Claire L.M.; Lanman, Brian A.; Liu, Longbin; Nishimura, Nobuko; Pettus, Liping H.; Reed, Anthony B.; Tadesse, Seifu; Tamayo, Nuria A.; Wurz, Ryan P.; Yang, Kevin; Andrews, Kristin L.; Whittington, Douglas A.; McCarter, John D.; Miguel, Tisha San; Zalameda, Leeanne; Jiang, Jian; Subramanian, Raju; Mullady, Erin L.; Caenepeel, Sean; Freeman, Daniel J.; Wang, Ling; Zhang, Nancy; Wu, Tian; Hughes, Paul E.; Norman, Mark H.

    2012-09-17

    A highly selective series of inhibitors of the class I phosphatidylinositol 3-kinases (PI3Ks) has been designed and synthesized. Starting from the dual PI3K/mTOR inhibitor 5, a structure-based approach was used to improve potency and selectivity, resulting in the identification of 54 as a potent inhibitor of the class I PI3Ks with excellent selectivity over mTOR, related phosphatidylinositol kinases, and a broad panel of protein kinases. Compound 54 demonstrated a robust PD-PK relationship inhibiting the PI3K/Akt pathway in vivo in a mouse model, and it potently inhibited tumor growth in a U-87 MG xenograft model with an activated PI3K/Akt pathway.

  5. Inhibition of Human Class I vs Class III Phosphatidylinositol 3'-Kinases.

    PubMed

    Hassett, Matthew R; Sternberg, Anna R; Roepe, Paul D

    2017-08-22

    Most investigations of phosphatidylinositol 3'-kinase (PI3K) drug inhibition have been via assays based on ADP appearance or ATP consumption (e.g., Liu, Q., et al. ( 2011 ) J. Med. Chem. 54 , 1473 - 1480 ). However, at least some PI3K isoforms show basal ATPase activity in the absence of PI lipid substrate(s), which may complicate quantification of drug potency, isoform specificity of some drugs, and synergy for drug combinations. In this study, we probe the class I vs class III isoform specificity of a selected set of PI3K inhibitors using a simple, inexpensive, semi high-throughput assay that quantifies production of phosphatidylinositol 3'-phosphate (PI3P) from phosphatidylinositol. Results are compared to previous data largely generated using ATPase activity assays. Good agreement between EC50 values computed via ATPase assays vs the reported PI3P formation assay is found for most drugs, but with a few exceptions. Furthermore, for the first time, drug inhibition of class I vs class III enzymes is compared side-by-side with the same assay for the important class I-specific inhibitors GSK2126458 ("Omipalisib") and NVP-BGT226 ("BGT226") currently in clinical development for advanced solid tumors.

  6. Cross-species Proteomics Reveals Specific Modulation of Signaling in Cancer and Stromal Cells by Phosphoinositide 3-kinase (PI3K) Inhibitors*

    PubMed Central

    Rajeeve, Vinothini; Vendrell, Iolanda; Wilkes, Edmund; Torbett, Neil; Cutillas, Pedro R.

    2014-01-01

    The tumor microenvironment plays key roles in cancer biology, but its impact on the regulation of signaling pathway activity in cancer cells has not been systemically investigated. We designed an analytical strategy that allows differential analysis of signaling between cancer and stromal cells present in tumor xenografts. We used this approach to investigate how in vivo growth conditions and PI3K inhibitors regulate pathway activities in both cancer and stromal cell populations. We found that, despite inducing more modest changes in protein expression, in vivo growing conditions extensively rewired protein kinase networks in cancer cells. As a result, different sets of phosphorylation sites were modulated by PI3K inhibitors in cancer cells growing in tumors relative to when these cells were in culture. The p110δ PI3K-selective compound CAL-101 (Idelalisib) did not inhibit markers of PI3K activity in cancer or stromal cells; however, unexpectedly, it induced phosphorylation on SQ motifs in both subpopulations of tumor cells in vivo but not in vitro. Thus, the interaction between cancer cells and the stroma modulated the ability of PI3K inhibitors to induce the activation of apoptosis in solid tumors. Our study provides proof-of-principle of a proteomics workflow for measuring signaling specifically in cancer and stromal cells and for investigating how cancer biochemistry is modulated in vivo. PMID:24648465

  7. Supramolecular nanoparticles that target phosphatidylinositol-3-kinase overcome insulin resistance and exert pronounced antitumor efficacy

    PubMed Central

    Kulkarni, Ashish A.; Roy, Bhaskar; Rao, Poornima S.; Wyant, Gregory A.; Mahmoud, Ayaat; Ramachandran, Madhumitha; Sengupta, Poulomi; Goldman, Aaron; Kotamraju, Venkata Ramana; Basu, Sudipta; Mashelkar, Raghunath A; Ruoslahti, Erkki; Dinulescu, Daniela M.; Sengupta, Shiladitya

    2013-01-01

    The centrality of phosphatidylinositol-3-kinase (PI3K) in cancer etiology is well established, but clinical translation of PI3K inhibitors has been limited by feedback signaling, suboptimal intra-tumoral concentration and an insulin resistance ‘class effect’. The current study was designed to explore the use of supramolecular nanochemistry for targeting PI3K to enhance antitumor efficacy and potentially overcome these limitations. PI3K inhibitor structures were rationally modified using a cholesterol-based derivative, facilitating supramolecular nanoassembly with L-α-phosphatidylcholine and DSPE-PEG. The supramolecular nanoparticles that were assembled were physicochemically characterized and functionally evaluated in vitro. Antitumor efficacy was quantified in vivo using 4T1 breast cancer and K-RasLSL/+/Ptenfl/fl ovarian cancer models, with effects on glucose homeostasis evaluated using an insulin sensitivity test. The use of PI103 and PI828 as surrogate molecules to engineer the supramolecular nanoparticles highlighted the need to keep design principles in perspective; specifically, potency of the active molecule and the linker chemistry were critical principles for efficacy, similar to antibody-drug conjugates. We found that the supramolecular nanoparticles exerted a temporally-sustained inhibition of phosphorylation of Akt, mTOR, S6K and 4EBP in vivo. These effects were associated with increased antitumor efficacy and survival as compared with PI103 and PI828. Efficacy was further increased by decorating the nanoparticle surface with tumor-homing peptides. Notably, the use of supramolecular nanoparticles abrogated the insulin resistance that has been associated widely with other PI3K inhibitors. This study provides a preclinical foundation for the use of supramolecular nanochemistry to overcome current challenges associated with PI3K inhibitors, offering a paradigm for extension to other molecularly targeted therapeutics being explored for cancer treatment

  8. Par3 integrates Tiam1 and phosphatidylinositol 3-kinase signaling to change apical membrane identity.

    PubMed

    Ruch, Travis R; Bryant, David M; Mostov, Keith E; Engel, Joanne N

    2017-01-15

    Pathogens can alter epithelial polarity by recruiting polarity proteins to the apical membrane, but how a change in protein localization is linked to polarity disruption is not clear. In this study, we used chemically induced dimerization to rapidly relocalize proteins from the cytosol to the apical surface. We demonstrate that forced apical localization of Par3, which is normally restricted to tight junctions, is sufficient to alter apical membrane identity through its interactions with phosphatidylinositol 3-kinase (PI3K) and the Rac1 guanine nucleotide exchange factor Tiam1. We further show that PI3K activity is required upstream of Rac1, and that simultaneously targeting PI3K and Tiam1 to the apical membrane has a synergistic effect on membrane remodeling. Thus, Par3 coordinates the action of PI3K and Tiam1 to define membrane identity, revealing a signaling mechanism that can be exploited by human mucosal pathogens. © 2017 Ruch et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  9. RhoG regulates anoikis through a phosphatidylinositol 3-kinase-dependent mechanism

    SciTech Connect

    Yamaki, Nao; Negishi, Manabu; Katoh, Hironori . E-mail: hirokato@pharm.kyoto-u.ac.jp

    2007-08-01

    In normal epithelial cells, cell-matrix interaction is required for cell survival and proliferation, whereas disruption of this interaction causes epithelial cells to undergo apoptosis called anoikis. Here we show that the small GTPase RhoG plays an important role in the regulation of anoikis. HeLa cells are capable of anchorage-independent cell growth and acquire resistance to anoikis. We found that RNA interference-mediated knockdown of RhoG promoted anoikis in HeLa cells. Previous studies have shown that RhoG activates Rac1 and induces several cellular functions including promotion of cell migration through its effector ELMO and the ELMO-binding protein Dock180 that function as a Rac-specific guanine nucleotide exchange factor. However, RhoG-induced suppression of anoikis was independent of the ELMO- and Dock180-mediated activation of Rac1. On the other hand, the regulation of anoikis by RhoG required phosphatidylinositol 3-kinase (PI3K) activity, and constitutively active RhoG bound to the PI3K regulatory subunit p85{alpha} and induced the PI3K-dependent phosphorylation of Akt. Taken together, these results suggest that RhoG protects cells from apoptosis caused by the loss of anchorage through a PI3K-dependent mechanism, independent of its activation of Rac1.

  10. Polycystin-1 Induces Resistance to Apoptosis through the Phosphatidylinositol 3-Kinase/Akt Signaling Pathway

    PubMed Central

    Boca, Manila; Distefano, Gianfranco; Boletta, Alessandra; Qian, Feng; Bhunia, Anil K.; Germino, Gregory G.

    2006-01-01

    Polycystin-1 (PC-1), the PKD1 gene product, is a large receptor whose expression in renal epithelial cells results in resistance to apoptosis and tubulogenesis, a model consistent with the phenotype observed in patients. This study links PC-1 expression to a signaling pathway that is known to be both antiapoptotic and important for normal tubulogenesis. This study found that PC-1 expression results in phosphorylation of Akt and downstream effectors and that phosphatidylinositol 3-kinase (PI3-K) inhibitors prevent this process. In addition, it is shown that dominant negative Akt can revert PC-1-induced protection from apoptosis. Furthermore, it was observed that increased PI3-K β activity in PC-1- expressing MDCK cells seems to be dependent on both tyrosine-kinase activity and heterotrimeric G proteins. It also was found that PC-1-induced tubulogenesis is inhibited by PI3-K inhibitors. Taken together, these data suggest that the PI3-K/Akt cascade may be a central modulator of PC-1 function and that its deregulation might be important in autosomal dominant polycystic kidney disease. PMID:16452497

  11. Berberine Induced Apoptosis of Human Osteosarcoma Cells by Inhibiting Phosphoinositide 3 Kinase/Protein Kinase B (PI3K/Akt) Signal Pathway Activation.

    PubMed

    Chen, Zhi-Ze

    2016-05-01

    Osteosarcoma is a malignant tumor with high mortality but effective therapy has not yet been developed. Berberine, an isoquinoline alkaloid component in several Chinese herbs including Huanglian, has been shown to induce growth inhibition and the apoptosis of certain cancer cells. The aim of this study was to determine the role of berberine on human osteosarcoma cell lines U2OS and its potential mechanism. The proliferation effect of U20S was exanimed by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di- phenytetrazoliumromide (MTT) and the percentage of apoptotic cells were determined by flow cytometric analysis. The expression of PI3K, p-Akt, Bax, Bcl-2, cleavage-PARP and Caspase3 were detected by Western blott. Berberine treatment caused dose-dependent inhibiting proliferation and inducing apoptosis of U20S cell. Mechanistically, berberine inhibits PI3K/AKT activation that, in turn, results in up-regulating the expression of Bax, and PARP and down-regulating the expression of Bcl-2 and caspase3. In all, berberine can suppress the proliferation and induce the apoptosis of U2OS cell through inhibiting the PI3K/Akt signaling pathway activation. Berberine can suppress the proliferation and induce the apoptosis of U2OS cell through inhibiting the PI3K/Akt signaling pathway activation.

  12. Berberine Induced Apoptosis of Human Osteosarcoma Cells by Inhibiting Phosphoinositide 3 Kinase/Protein Kinase B (PI3K/Akt) Signal Pathway Activation

    PubMed Central

    2016-01-01

    Background: Osteosarcoma is a malignant tumor with high mortality but effective therapy has not yet been developed. Berberine, an isoquinoline alkaloid component in several Chinese herbs including Huanglian, has been shown to induce growth inhibition and the apoptosis of certain cancer cells. The aim of this study was to determine the role of berberine on human osteosarcoma cell lines U2OS and its potential mechanism. Methods: The proliferation effect of U20S was exanimed by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di- phenytetrazoliumromide (MTT) and the percentage of apoptotic cells were determined by flow cytometric analysis. The expression of PI3K, p-Akt, Bax, Bcl-2, cleavage-PARP and Caspase3 were detected by Western blott. Results: Berberine treatment caused dose-dependent inhibiting proliferation and inducing apoptosis of U20S cell. Mechanistically, berberine inhibits PI3K/AKT activation that, in turn, results in up-regulating the expression of Bax, and PARP and down-regulating the expression of Bcl-2 and caspase3. In all, berberine can suppress the proliferation and induce the apoptosis of U2OS cell through inhibiting the PI3K/Akt signaling pathway activation. Conclusion: Berberine can suppress the proliferation and induce the apoptosis of U2OS cell through inhibiting the PI3K/Akt signaling pathway activation. PMID:27398330

  13. The phosphatidylinositol 3-kinase/Akt/mTOR signaling network as a therapeutic target in acute myelogenous leukemia patients

    PubMed Central

    Martelli, Alberto M.; Evangelisti, Camilla; Chiarini, Francesca; McCubrey, James A.

    2010-01-01

    The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling axis plays a central role in cell proliferation, growth, and survival under physiological conditions. However, aberrant PI3K/Akt/mTOR signaling has been implicated in many human cancers, including acute myelogenous leukemia (AML). Therefore, the PI3K/Akt/mTOR network is considered as a validated target for innovative cancer therapy. The limit of acceptable toxicity for standard polychemotherapy has been reached in AML. Novel therapeutic strategies are therefore needed. This review highlights how the PI3K/Akt/mTOR signaling axis is constitutively active in AML patients, where it affects survival, proliferation, and drug-resistance of leukemic cells including leukemic stem cells. Effective targeting of this pathway with small molecule kinase inhibitors, employed alone or in combination with other drugs, could result in the suppression of leukemic cell growth. Furthermore, targeting the PI3K/Akt/mTOR signaling network with small pharmacological inhibitors, employed either alone or in combinations with other drugs, may result in less toxic and more efficacious treatment of AML patients. Efforts to exploit pharmacological inhibitors of the PI3K/Akt/mTOR cascade which show efficacy and safety in the clinical setting are now underway. PMID:20671809

  14. Involvement of Phosphatidylinositol 3-kinase in the regulation of proline catabolism in Arabidopsis thaliana

    PubMed Central

    Leprince, Anne-Sophie; Magalhaes, Nelly; De Vos, Delphine; Bordenave, Marianne; Crilat, Emilie; Clément, Gilles; Meyer, Christian; Munnik, Teun; Savouré, Arnould

    2015-01-01

    Plant adaptation to abiotic stresses such as drought and salinity involves complex regulatory processes. Deciphering the signaling components that are involved in stress signal transduction and cellular responses is of importance to understand how plants cope with salt stress. Accumulation of osmolytes such as proline is considered to participate in the osmotic adjustment of plant cells to salinity. Proline accumulation results from a tight regulation between its biosynthesis and catabolism. Lipid signal components such as phospholipases C and D have previously been shown to be involved in the regulation of proline metabolism in Arabidopsis thaliana. In this study, we demonstrate that proline metabolism is also regulated by class-III Phosphatidylinositol 3-kinase (PI3K), VPS34, which catalyses the formation of phosphatidylinositol 3-phosphate (PI3P) from phosphatidylinositol. Using pharmacological and biochemical approaches, we show that the PI3K inhibitor, LY294002, affects PI3P levels in vivo and that it triggers a decrease in proline accumulation in response to salt treatment of A. thaliana seedlings. The lower proline accumulation is correlated with a lower transcript level of Pyrroline-5-carboxylate synthetase 1 (P5CS1) biosynthetic enzyme and higher transcript and protein levels of Proline dehydrogenase 1 (ProDH1), a key-enzyme in proline catabolism. We also found that the ProDH1 expression is induced in a pi3k-hemizygous mutant, further demonstrating that PI3K is involved in the regulation of proline catabolism through transcriptional regulation of ProDH1. A broader metabolomic analysis indicates that LY294002 also reduced other metabolites, such as hydrophobic and aromatic amino acids and sugars like raffinose. PMID:25628629

  15. Genetic alterations in the phosphatidylinositol-3 kinase/Akt pathway in thyroid cancer.

    PubMed

    Xing, Mingzhao

    2010-07-01

    Aberrant activation of the phosphatidylinositol-3 kinase (PI3K)/Akt pathway plays a fundamental role in thyroid tumorigenesis, particularly in follicular thyroid cancer (FTC) and aggressive thyroid cancer, such as anaplastic thyroid cancer (ATC). As the drivers of this process, many genetic alterations activating the PI3K/Akt pathway have been identified in thyroid cancer in recent years. This review summarizes the current knowledge on major genetic alterations in the PI3K/Akt pathway. These include PIK3CA mutations and genomic amplification/copy gain, Ras mutations, PTEN mutations, RET/PTC and PPARgamma/Pax8 rearrangements, as well as amplification/copy gain of PIK3CB, PDK1, Akt, and various receptor tyrosine kinase genes. Most of these genetic alterations are particularly common in FTC and many of them are even more common in ATC; they are generally less common in papillary thyroid cancer (PTC), in which the MAP kinase (MAPK) pathway activated by the BRAF mutation instead plays a major role. Methylation and, thus, epigenetic silencing of PTEN, a major negative regulator of the PI3K/Akt pathway, occurs in close association with activating genetic alterations of the PI3K/Akt pathway, constituting a unique self-enhancement mechanism for this pathway. Many of these genetic alterations are mutually exclusive in differentiated thyroid tumors, but with increasing concurrence from benign tumors to FTC to ATC. RET/PTC, Ras, and receptor tyrosine kinase could dually activate the PI3K/Akt and MAPK pathways. Most cases of ATC harbor genetic alterations in these genes or other genetic combinations that can activate both pathways. It is proposed that genetic alterations in the PI3K/Akt pathway promote thyroid cell transformation to FTC and that genetic alterations in the MAPK pathway promote cell transformation to PTC; accumulation of multiple genetic alterations that can activate both pathways promotes thyroid cancer aggressiveness and progression to ATC. Genetic alterations

  16. Genetic Alterations in the Phosphatidylinositol-3 Kinase/Akt Pathway in Thyroid Cancer

    PubMed Central

    2010-01-01

    Background Aberrant activation of the phosphatidylinositol-3 kinase (PI3K)/Akt pathway plays a fundamental role in thyroid tumorigenesis, particularly in follicular thyroid cancer (FTC) and aggressive thyroid cancer, such as anaplastic thyroid cancer (ATC). As the drivers of this process, many genetic alterations activating the PI3K/Akt pathway have been identified in thyroid cancer in recent years. Summary This review summarizes the current knowledge on major genetic alterations in the PI3K/Akt pathway. These include PIK3CA mutations and genomic amplification/copy gain, Ras mutations, PTEN mutations, RET/PTC and PPARγ/Pax8 rearrangements, as well as amplification/copy gain of PIK3CB, PDK1, Akt, and various receptor tyrosine kinase genes. Most of these genetic alterations are particularly common in FTC and many of them are even more common in ATC; they are generally less common in papillary thyroid cancer (PTC), in which the MAP kinase (MAPK) pathway activated by the BRAF mutation instead plays a major role. Methylation and, thus, epigenetic silencing of PTEN, a major negative regulator of the PI3K/Akt pathway, occurs in close association with activating genetic alterations of the PI3K/Akt pathway, constituting a unique self-enhancement mechanism for this pathway. Many of these genetic alterations are mutually exclusive in differentiated thyroid tumors, but with increasing concurrence from benign tumors to FTC to ATC. RET/PTC, Ras, and receptor tyrosine kinase could dually activate the PI3K/Akt and MAPK pathways. Most cases of ATC harbor genetic alterations in these genes or other genetic combinations that can activate both pathways. It is proposed that genetic alterations in the PI3K/Akt pathway promote thyroid cell transformation to FTC and that genetic alterations in the MAPK pathway promote cell transformation to PTC; accumulation of multiple genetic alterations that can activate both pathways promotes thyroid cancer aggressiveness and progression to ATC

  17. Phosphatidylinositol 3-kinase-dependent signaling modulates taurochenodeoxycholic acid-induced liver injury and cholestasis in perfused rat livers.

    PubMed

    Rust, Christian; Bauchmuller, Kris; Fickert, Peter; Fuchsbichler, Andrea; Beuers, Ulrich

    2005-07-01

    Taurochenodeoxycholic acid (TCDCA), but not glycochenodeoxycholic acid (GCDCA), activates a phosphatidylinositol 3-kinase (PI3-K)-mediated survival pathway in vitro. Here, the effects of PI3-K inhibition on TCDCA- and GCDCA-induced hepatocellular injury, apoptosis, and bile secretion were examined in the intact liver. In isolated perfused rat livers, bile flow was determined gravimetrically. Hepatovenous lactate dehydrogenase and alanine aminotransferase efflux as markers of liver integrity and biliary secretion of 2,4-dinitrophenyl-S-glutathione (DNP-GS) were determined photometrically. Apoptosis was assessed by immunohistochemistry of active caspase-3 and cytokeratin 18 in liver tissue. Phosphorylation of protein kinase B (PKB/Akt) as a readout of PI3-K activity was determined by immunoblot analysis. Bile acid concentrations were determined by gas chromatography. TCDCA (25 muM) induced moderate liver injury by hepatocellular apoptosis and distinctly reduced bile flow and DNP-GS secretion. In contrast, GCDCA (25 muM) induced severe liver injury by extensive hepatocyte apoptosis. TCDCA strongly activated PI3-K, whereas GCDCA did not markedly affect PI3-K activity. Inhibition of PI3-K by 100 nM wortmannin enhanced TCDCA-induced liver injury and apoptosis and tended to aggravate the cholestatic effect of TCDCA. In contrast, wortmannin reduced GCDCA-induced liver injury and apoptosis. Bile acid uptake tended to be reduced by wortmannin. The cholestatic effect of GCDCA was aggravated by wortmannin. Inhibition of PI3-K markedly aggravated TCDCA-induced but not GCDCA-induced liver damage and hepatocyte apoptosis. Thus TCDCA appears to block its inherent toxicity by a PI3-K-dependent survival pathway in the intact liver.

  18. Phosphatidylinositol 3-Kinase Plays a Vital Role in Regulation of Rice Seed Vigor via Altering NADPH Oxidase Activity

    PubMed Central

    Liu, Jian; Zhou, Jun; Xing, Da

    2012-01-01

    Phosphatidylinositol 3-kinase (PI3K) has been reported to be important in normal plant growth and stress responses. In this study, it was verified that PI3K played a vital role in rice seed germination through regulating NADPH oxidase activity. Suppression of PI3K activity by inhibitors wortmannin or LY294002 could abate the reactive oxygen species (ROS) formation, which resulted in disturbance to the seed germination. And then, the signal cascades that PI3K promoted the ROS liberation was also evaluated. Diphenylene iodonium (DPI), an NADPH oxidase inhibitor, suppressed most of ROS generation in rice seed germination, which suggested that NADPH oxidase was the main source of ROS in this process. Pharmacological experiment and RT-PCR demonstrated that PI3K promoted the expression of Os rboh9. Moreover, functional analysis by native PAGE and the measurement of the 2, 3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazo-lium-5- carboxanilide (XTT) formazan concentration both showed that PI3K promoted the activity of NADPH oxidase. Furthermore, the western blot analysis of OsRac-1 demonstrated that the translocation of Rac-1 from cytoplasm to plasma membrane, which was known as a key factor in the assembly of NADPH oxidase, was suppressed by treatment with PI3K inhibitors, resulting in the decreased activity of NADPH oxidase. Taken together, these data favored the novel conclusion that PI3K regulated NADPH oxidase activity through modulating the recruitment of Rac-1 to plasma membrane and accelerated the process of rice seed germination. PMID:22448275

  19. LY294002 inhibits glucocorticoid-induced COX-2 gene expression in cardiomyocytes through a phosphatidylinositol 3 kinase-independent mechanism

    SciTech Connect

    Sun Haipeng; Xu Beibei; Sheveleva, Elena; Chen, Qin M.

    2008-10-01

    Glucocorticoids induce COX-2 expression in rat cardiomyocytes. While investigating whether phosphatidylinositol 3 kinase (PI3K) plays a role in corticosterone (CT)-induced COX-2, we found that LY294002 (LY29) but not wortmannin (WM) attenuates CT from inducing COX-2 gene expression. Expression of a dominant-negative mutant of p85 subunit of PI3K failed to inhibit CT from inducing COX-2 expression. CT did not activate PI3K/AKT signaling pathway whereas LY29 and WM decreased the activity of PI3K. LY303511 (LY30), a structural analogue and a negative control for PI3K inhibitory activity of LY29, also suppressed COX-2 induction. These data suggest PI3K-independent mechanisms in regulating CT-induced COX-2 expression. LY29 and LY30 do not inhibit glucocorticoid receptor transactivity. Both compounds have been reported to inhibit Casein Kinase 2 activity and modulate potassium and calcium levels independent of PI3K, while LY29 has been reported to inhibit mammalian Target of Rapamycin (mTOR), and DNA-dependent Protein Kinase (DNA-PK). Inhibitor of Casein Kinase 2 (CK2), mTOR or DNA-PK failed to prevent CT from inducing COX-2 expression. Tetraethylammonium (TEA), a potassium channel blocker, and nimodipine, a calcium channel blocker, both attenuated CT from inducing COX-2 gene expression. CT was found to increase intracellular Ca{sup 2+} concentration, which can be inhibited by LY29, TEA or nimodipine. These data suggest a possible role of calcium instead of PI3K in CT-induced COX-2 expression in cardiomyocytes.

  20. Phosphatidylinositol 3-kinase plays a vital role in regulation of rice seed vigor via altering NADPH oxidase activity.

    PubMed

    Liu, Jian; Zhou, Jun; Xing, Da

    2012-01-01

    Phosphatidylinositol 3-kinase (PI3K) has been reported to be important in normal plant growth and stress responses. In this study, it was verified that PI3K played a vital role in rice seed germination through regulating NADPH oxidase activity. Suppression of PI3K activity by inhibitors wortmannin or LY294002 could abate the reactive oxygen species (ROS) formation, which resulted in disturbance to the seed germination. And then, the signal cascades that PI3K promoted the ROS liberation was also evaluated. Diphenylene iodonium (DPI), an NADPH oxidase inhibitor, suppressed most of ROS generation in rice seed germination, which suggested that NADPH oxidase was the main source of ROS in this process. Pharmacological experiment and RT-PCR demonstrated that PI3K promoted the expression of Os rboh9. Moreover, functional analysis by native PAGE and the measurement of the 2, 3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazo-lium-5- carboxanilide (XTT) formazan concentration both showed that PI3K promoted the activity of NADPH oxidase. Furthermore, the western blot analysis of OsRac-1 demonstrated that the translocation of Rac-1 from cytoplasm to plasma membrane, which was known as a key factor in the assembly of NADPH oxidase, was suppressed by treatment with PI3K inhibitors, resulting in the decreased activity of NADPH oxidase. Taken together, these data favored the novel conclusion that PI3K regulated NADPH oxidase activity through modulating the recruitment of Rac-1 to plasma membrane and accelerated the process of rice seed germination.

  1. Phosphoinositide 3-Kinase (PI3K) Subunit p110δ Is Essential for Trophoblast Cell Differentiation and Placental Development in Mouse

    PubMed Central

    Hu, Xiwen; Li, Jiangchao; Zhang, Qianqian; Zheng, Lingyun; Wang, Guang; Zhang, Xiaohan; Zhang, Jingli; Gu, Quliang; Ye, Yuxiang; Guo, Sun-Wei; Yang, Xuesong; Wang, Lijing

    2016-01-01

    Maternal PI3K p110δ has been implicated in smaller litter sizes in mice, but its underlying mechanism remains unclear. The placenta is an indispensable chimeric organ that supports mammalian embryonic development. Using a mouse model of genetic inactivation of PI3K p110δ (p110δD910A/D910A), we show that fetuses carried by p110δD910A/D910A females were growth retarded and showed increased mortality in utero mainly during placentation. The placentas in p110δD910A/D910A females were anomalously anemic, exhibited thinner spongiotrophoblast layer and looser labyrinth zone, which indicate defective placental vasculogenesis. In addition, p110δ was detected in primary trophoblast giant cells (P-TGC) at early placentation. Maternal PI3K p110δ inactivation affected normal TGCs generation and expansion, impeded the branching of chorioallantoic placenta but enhanced the expression of matrix metalloproteinases (MMP-2, MMP-12). Poor vasculature support for the developing fetoplacental unit resulted in fetal death or gross growth retardation. These data, taken together, provide the first in vivo evidence that p110δ may play an important role in placental vascularization through manipulating trophoblast giant cell. PMID:27306493

  2. Phosphatidylinositol 3-kinase and NF-kappa B/Rel are at the divergence of CD40-mediated proliferation and survival pathways.

    PubMed

    Andjelic, S; Hsia, C; Suzuki, H; Kadowaki, T; Koyasu, S; Liou, H C

    2000-10-01

    CD40 receptor ligation evokes several crucial outcomes for the fate of an activated B cell, including proliferation and survival. Although multiple signaling molecules in the CD40 pathways have been identified, their specific roles in regulating proliferation and maintaining cell viability are still obscure. In this report, we demonstrate that the activation of both phosphatidylinositol 3-kinase (PI-3K) and NF-kappaB/Rel transcription factors is crucial for CD40-mediated proliferation. Furthermore, our data indicate that PI-3K is indispensable for CD40-mediated NF-kappaB/Rel activation. This is achieved via activation of AKT and the degradation of IkappaBalpha. Furthermore, we show that PI-3K activity is necessary for the degradation of cyclin-dependent kinase inhibitor p27kip. Therefore, both of these events comprise the mechanism by which PI-3K controls cell proliferation. In contrast to the absolute requirement of PI-3K and NF-kappaB/Rel for proliferation, these signaling molecules are only partially responsible for CD40-mediated survival, as blocking of PI-3K activity did not lead to apoptosis of anti-CD40-treated cells. However, the PI-3K/NF-kappaB pathway is still required for CD40-induced Bcl-X gene expression. Taken together, our data indicate that multiple survival pathways are triggered via this receptor, whereas NF-kappaB/Rel and PI-3K are crucial for CD40-induced proliferation.

  3. Drugging PI3K in cancer: refining targets and therapeutic strategies

    PubMed Central

    Yap, Timothy A; Bjerke, Lynn; Clarke, Paul A; Workman, Paul

    2015-01-01

    The phosphatidylinositol-3 kinase (PI3K) pathway is one of the most frequently activated pathogenic signalling routes in human cancers, making it a rational and important target for innovative anticancer drug development and precision medicine. The three main classes of PI3K inhibitors currently in clinical testing comprise dual pan-Class I PI3K/mTOR inhibitors, pan-Class I PI3K inhibitors lacking significant mTOR activity and isoform-selective PI3K inhibitors. A major step forward in recent years is the progression of over 30 small molecule PI3K inhibitors into clinical trials and the first regulatory approval of the PI3Kδ inhibitor idelalisib for multiple B-cell malignancies. This review article focuses on the progress made in the discovery and development of novel PI3K inhibitors, with an emphasis on antitumour activity and tolerability profiles for agents that have entered clinical trials. We also discuss the key issues of drug resistance, patient selection approaches and rational targeted combinations. Finally, we envision the future development and use of PI3K inhibitors for the treatment of patients with a range of malignancies. PMID:26117819

  4. Differential sensitivities of trastuzumab (Herceptin)-resistant human breast cancer cells to phosphoinositide-3 kinase (PI-3K) and epidermal growth factor receptor (EGFR) kinase inhibitors.

    PubMed

    Chan, Carmel T; Metz, Marianne Z; Kane, Susan E

    2005-05-01

    Her2 (erbB2/neu) is overexpressed in 25-30% of human breast cancers. Herceptin is a recombinant humanized Her2 antibody used to treat breast cancer patients with Her2 overexpression. Over a 5-month selection process, we isolated clones of BT474 (BT) human breast carcinoma cells (BT/Her(R)) that were resistant to Herceptin in vitro. In BT/Her(R) subclones, cell-surface, phosphorylated and total cellular Her2 protein remained high in the continuous presence of Herceptin. Likewise, the levels of cell-surface, phosphorylated, and total cellular Her3 and EGFR were either unchanged or only slightly elevated in BT/Her(R) subclones relative to BT cells. One BT/Her(R) subclone had substantially upregulated cell-surface EGFR, but this did not correlate with a higher relative resistance to Herceptin. In looking at the downstream PI-3K/Akt signaling pathway, phosphorylated and total Akt levels and Akt kinase activities were all sustained in BT/Her(R) subclones in the presence of Herceptin, but significantly downregulated in BT cells exposed to Herceptin. Whereas BT cells lost sensitivity to the PI-3K inhibitor LY294002 in the presence of Herceptin, BT/Her(R) subclones were equally sensitive to this agent in the presence and absence of Herceptin. This suggests that BT/Her(R) subclones acquired a Herceptin-resistant mechanism of PI-3K signaling. BT/Her(R) subclones were also sensitive to the EGFR kinase inhibitor AG1478 in the presence of Herceptin, to the same extent as BT cells. The BT/Her(R) subclones provide new insights into mechanisms of Herceptin resistance and suggest new treatment strategies in combination with other inhibitors targeted to signal transduction pathways.

  5. Inhibition of phosphatidylinositol 3-kinase stimulates activity of the small-conductance K channel in the CCD

    PubMed Central

    Li, Dimin; Wei, Yuan; Babilonia, Elisa; Wang, Zhijian; Wang, Wen-Hui

    2010-01-01

    We used Western blotting to examine the expression of phosphatidylinositol 3-kinase (PI3K) in the renal cortex and outer medulla and employed the patch-clamp technique to study the effect of PI3K on the ROMK-like small-conductance K (SK) channels in the cortical collecting duct (CCD). Low K intake increased the expression of the 110-kDa α-subunit (p110α) of PI3K compared with rats on a normal-K diet. Because low K intake increases superoxide levels (2), the possibility that increases in superoxide anions may be responsible for the effect of low K intake on the expression of PI3K is supported by finding that addition of H2O2 stimulates the expression of p110α in M1 cells. Inhibition of PI3K with either wortmannin or LY-294002 significantly increased channel activity in the CCD from rats on a K-deficient (KD) diet or on a normal-K diet. The stimulatory effect of wortmannin on ROMK channel activity cannot be mimicked by inhibition of phospholipase C with U-73122. This suggests that the effect of inhibiting PI3K was not the result of increasing the phosphatidylinositol 4,5-bisphosphate level. Moreover, application of the exogenous phosphatidylinositol 3,4,5-trisphosphate analog had no effect on channel activity in excised patches. Because low K intake has been shown to increase the activity of protein tyrosine kinase (PTK), we explored the role of the interaction between PTK and PI3K in the regulation of the SK channel activity. Inhibition of PTK increased SK channel activity in the CCD from rats on a KD diet. However, addition of wortmannin did not further increase ROMK channel activity. Also, the effect of wortmannin was abolished by treatment of CCD with phalloidin. We conclude that PI3K is involved in mediating the effect of low K intake on ROMK channel activity in the CCD and that the effect of PI3K on SK channels requires the involvement of PTK and the cytoskeleton. PMID:16204406

  6. Inhibition of phosphatidylinositol 3-kinase stimulates activity of the small-conductance K channel in the CCD.

    PubMed

    Li, Dimin; Wei, Yuan; Babilonia, Elisa; Wang, Zhijian; Wang, Wen-Hui

    2006-04-01

    We used Western blotting to examine the expression of phosphatidylinositol 3-kinase (PI3K) in the renal cortex and outer medulla and employed the patch-clamp technique to study the effect of PI3K on the ROMK-like small-conductance K (SK) channels in the cortical collecting duct (CCD). Low K intake increased the expression of the 110-kDa alpha-subunit (p110alpha) of PI3K compared with rats on a normal-K diet. Because low K intake increases superoxide levels (2), the possibility that increases in superoxide anions may be responsible for the effect of low K intake on the expression of PI3K is supported by finding that addition of H(2)O(2) stimulates the expression of p110alpha in M1 cells. Inhibition of PI3K with either wortmannin or LY-294002 significantly increased channel activity in the CCD from rats on a K-deficient (KD) diet or on a normal-K diet. The stimulatory effect of wortmannin on ROMK channel activity cannot be mimicked by inhibition of phospholipase C with U-73122. This suggests that the effect of inhibiting PI3K was not the result of increasing the phosphatidylinositol 4,5-bisphosphate level. Moreover, application of the exogenous phosphatidylinositol 3,4,5-trisphosphate analog had no effect on channel activity in excised patches. Because low K intake has been shown to increase the activity of protein tyrosine kinase (PTK), we explored the role of the interaction between PTK and PI3K in the regulation of the SK channel activity. Inhibition of PTK increased SK channel activity in the CCD from rats on a KD diet. However, addition of wortmannin did not further increase ROMK channel activity. Also, the effect of wortmannin was abolished by treatment of CCD with phalloidin. We conclude that PI3K is involved in mediating the effect of low K intake on ROMK channel activity in the CCD and that the effect of PI3K on SK channels requires the involvement of PTK and the cytoskeleton.

  7. Phosphatidylinositol 3-kinase signaling determines kidney size.

    PubMed

    Chen, Jian-Kang; Nagai, Kojiro; Chen, Jianchun; Plieth, David; Hino, Masayo; Xu, Jinxian; Sha, Feng; Ikizler, T Alp; Quarles, C Chad; Threadgill, David W; Neilson, Eric G; Harris, Raymond C

    2015-06-01

    Kidney size adaptively increases as mammals grow and in response to the loss of 1 kidney. It is not clear how kidneys size themselves or if the processes that adapt kidney mass to lean body mass also mediate renal hypertrophy following unilateral nephrectomy (UNX). Here, we demonstrated that mice harboring a proximal tubule-specific deletion of Pten (Pten(ptKO)) have greatly enlarged kidneys as the result of persistent activation of the class I PI3K/mTORC2/AKT pathway and an increase of the antiproliferative signals p21(Cip1/WAF) and p27(Kip1). Administration of rapamycin to Pten(ptKO) mice diminished hypertrophy. Proximal tubule-specific deletion of Egfr in Pten(ptKO) mice also attenuated class I PI3K/mTORC2/AKT signaling and reduced the size of enlarged kidneys. In Pten(ptKO) mice, UNX further increased mTORC1 activation and hypertrophy in the remaining kidney; however, mTORC2-dependent AKT phosphorylation did not increase further in the remaining kidney of Pten(ptKO) mice, nor was it induced in the remaining kidney of WT mice. After UNX, renal blood flow and amino acid delivery to the remaining kidney rose abruptly, followed by increased amino acid content and activation of a class III PI3K/mTORC1/S6K1 pathway. Thus, our findings demonstrate context-dependent roles for EGFR-modulated class I PI3K/mTORC2/AKT signaling in the normal adaptation of kidney size and PTEN-independent, nutrient-dependent class III PI3K/mTORC1/S6K1 signaling in the compensatory enlargement of the remaining kidney following UNX.

  8. Evolutionary history of phosphatidylinositol- 3-kinases: ancestral origin in eukaryotes and complex duplication patterns.

    PubMed

    Philippon, Héloïse; Brochier-Armanet, Céline; Perrière, Guy

    2015-10-19

    Phosphatidylinositol-3-kinases (PI3Ks) are a family of eukaryotic enzymes modifying phosphoinositides in phosphatidylinositols-3-phosphate. Located upstream of the AKT/mTOR signalling pathway, PI3Ks activate secondary messengers of extracellular signals. They are involved in many critical cellular processes such as cell survival, angiogenesis and autophagy. PI3K family is divided into three classes, including 14 human homologs. While class II enzymes are composed of a single catalytic subunit, class I and III also contain regulatory subunits. Here we present an in-depth phylogenetic analysis of all PI3K proteins. We confirmed that PI3K catalytic subunits form a monophyletic group, whereas regulatory subunits form three distinct groups. The phylogeny of the catalytic subunits indicates that they underwent two major duplications during their evolutionary history: the most ancient arose in the Last Eukaryotic Common Ancestor (LECA) and led to the emergence of class III and class I/II, while the second - that led to the separation between class I and II - occurred later, in the ancestor of Unikonta (i.e., the clade grouping Amoebozoa, Fungi, and Metazoa). These two major events were followed by many lineage specific duplications in particular in vertebrates, but also in various protist lineages. Major loss events were also detected in Vidiriplantae and Fungi. For the regulatory subunits, we identified homologs of class III in all eukaryotic groups indicating that, for this class, both the catalytic and the regulatory subunits were presents in LECA. In contrast, homologs of the regulatory class I have a more recent origin. The phylogenetic analysis of the PI3K shed a new light on the evolutionary history of these enzymes. We found that LECA already contained a PI3K class III composed of a catalytic and a regulatory subunit. Absence of class II regulatory subunits and the recent origin of class I regulatory subunits is puzzling given that the class I/II catalytic subunit

  9. Phosphatidylinositol 3-Kinase Mediates Bronchioalveolar Stem Cell Expansion in Mouse Models of Oncogenic K-ras-Induced Lung Cancer

    PubMed Central

    Yang, Yanan; Iwanaga, Kentaro; Raso, Maria Gabriela; Wislez, Marie; Hanna, Amy E.; Wieder, Eric D.; Molldrem, Jeffrey J.; Wistuba, Ignacio I.; Powis, Garth; Demayo, Francesco J.; Kim, Carla F.; Kurie, Jonathan M.

    2008-01-01

    Background Non-small cell lung cancer (NSCLC) is the most common cause of cancer-related death in Western countries. Developing more effective NSCLC therapeutics will require the elucidation of the genetic and biochemical bases for this disease. Bronchioalveolar stem cells (BASCs) are a putative cancer stem cell population in mouse models of oncogenic K-ras-induced lung adenocarcinoma, an histologic subtype of NSCLC. The signals activated by oncogenic K-ras that mediate BASC expansion have not been fully defined. Methodology/Principal Findings We used genetic and pharmacologic approaches to modulate the activity of phosphatidylinositol 3-kinase (PI3K), a key mediator of oncogenic K-ras, in two genetic mouse models of lung adenocarcinoma. Oncogenic K-ras-induced BASC accumulation and tumor growth were blocked by treatment with a small molecule PI3K inhibitor and enhanced by inactivation of phosphatase and tensin homologue deleted from chromosome 10, a negative regulator of PI3K. Conclusions/Significance We conclude that PI3K is a critical regulator of BASC expansion, supporting treatment strategies to target PI3K in NSCLC patients. PMID:18493606

  10. IPD-196, a novel phosphatidylinositol 3-kinase inhibitor with potent anticancer activity against hepatocellular carcinoma.

    PubMed

    Lee, Ju-Hee; Lee, Hyunseung; Yun, Sun-Mi; Jung, Kyung Hee; Jeong, Yujeong; Yan, Hong Hua; Hong, Sungwoo; Hong, Soon-Sun

    2013-02-01

    As the activation of phosphatidylinositol 3-kinase (PI3K) is associated with a wide variety of human malignancies, it is emerging as an attractive target for cancer treatment. In this study we synthesized a novel PI3Kα inhibitor, IPD-196 [ethyl 6-(5-(2,4-difluorophenylsulfonamido)pyridin-3-yl)imidazo[1,2-a]pyridine-3-carboxylate], and evaluated its anticancer effects on human hepatocellular carcinoma (HCC) cells. IPD-196 effectively inhibited the phosphorylation of downstream PI3K effectors such as Akt, mTOR, p70S6K, and 4E-BP1, and its antiproliferative effect was more potent than that of sorafenib or LY294002. It also induced cell cycle arrest at the G0/G1 phase as well as apoptosis by increasing the proportion of sub-G1 apoptotic cells, and the levels of cleaved PARP, caspase-3, and caspase-9. Furthermore, it decreased the expression of HIF-1α and VEGF in Huh-7 cells, and inhibited tube formation and migration of human umbilical vein endothelial cells, which was confirmed by a Matrigel plug assay in mice. Taken together, IPD-196 exhibited its anticancer activity through disruption of the PI3K/Akt pathway that caused cell cycle arrest, apoptosis induction, and inhibition of angiogenesis in human HCC cells. We therefore suggest that IPD-196 may be a potential candidate drug for targeted HCC therapy.

  11. Status of PI3K/Akt/mTOR Pathway Inhibitors in Lymphoma

    PubMed Central

    Westin, Jason R.

    2014-01-01

    The phosphatidylinositol-3-kinase (PI3K) pathway is well known to regulate a wide variety of essential cellular functions, including glucose metabolism, translational regulation of protein synthesis, cell proliferation, apoptosis, and survival. Aberrations in the PI3K pathway are among the most frequently observed in cancer, and include amplifications, rearrangements, mutations, and loss of regulators. As a net result of these anomalies, the PI3K pathway is activated in many malignancies, including in Hodgkin and non-Hodgkin lymphomas, and yields a competitive growth and survival advantage, increased metastatic ability, and resistance to conventional therapy. Numerous inhibitors targeting various nodes in the PI3K pathway are undergoing clinical development, and their current status in lymphoma will be the focus of this review. PMID:24650973

  12. PI3K Functions in Cancer Progression, Anticancer Immunity and Immune Evasion by Tumors

    PubMed Central

    Dituri, Francesco; Mazzocca, Antonio; Giannelli, Gianluigi; Antonaci, Salvatore

    2011-01-01

    The immunological surveillance of tumors relies on a specific recognition of cancer cells and their associate antigens by leucocytes of innate and adaptive immune responses. However, a dysregulated cytokine release can lead to, or be associated with, a failure in cell-cell recognition, thus, allowing cancer cells to evade the killing system. The phosphatidylinositol 3-kinase (PI3K) pathway regulates multiple cellular processes which underlie immune responses against pathogens or malignant cells. Conversely, there is accumulating evidence that the PI3K pathway is involved in the development of several malignant traits of cancer cells as well as their escape from immunity. Herein, we review the counteracting roles of PI3K not only in antitumor immune response but also in the mechanisms that cancer cells use to avoid leukocyte attack. In addition, we discuss, from antitumor immunological point of view, the potential benefits and disadvantages arising from use of anticancer pharmacological agents targeting the PI3K pathway. PMID:22046194

  13. Two hits are better than one: targeting both phosphatidylinositol 3-kinase and mammalian target of rapamycin as a therapeutic strategy for acute leukemia treatment

    PubMed Central

    Martelli, Alberto M.; Chiarini, Francesca; Evangelisti, Camilla; Cappellini, Alessandra; Buontempo, Francesca; Bressanin, Daniela; Fini, Milena; McCubrey, James A.

    2012-01-01

    Phosphatidylinositol 3-kinase (PI3K) and mammalian target of rapamycin (mTOR) are two key components of the PI3K/Akt/mTOR signaling pathway. This signal transduction cascade regulates a wide range of physiological cell processes, that include differentiation, proliferation, apoptosis, autophagy, metabolism, motility, and exocytosis. However, constitutively active PI3K/Akt/mTOR signaling characterizes many types of tumors where it negatively influences response to therapeutic treatments. Hence, targeting PI3K/Akt/mTOR signaling with small molecule inhibitors may improve cancer patient outcome. The PI3K/Akt/mTOR signaling cascade is overactive in acute leukemias, where it correlates with enhanced drug-resistance and poor prognosis. The catalytic sites of PI3K and mTOR share a high degree of sequence homology. This feature has allowed the synthesis of ATP-competitive compounds targeting the catalytic site of both kinases. In preclinical models, dual PI3K/mTOR inhibitors displayed a much stronger cytotoxicity against acute leukemia cells than either PI3K inhibitors or allosteric mTOR inhibitors, such as rapamycin. At variance with rapamycin, dual PI3K/mTOR inhibitors targeted both mTOR complex 1 and mTOR complex 2, and inhibited the rapamycin-resistant phosphorylation of eukaryotic initiation factor 4E-binding protein 1, resulting in a marked inhibition of oncogenic protein translation. Therefore, they strongly reduced cell proliferation and induced an important apoptotic response. Here, we reviewed the evidence documenting that dual PI3K/mTOR inhibitors may represent a promising option for future targeted therapies of acute leukemia patients. PMID:22564882

  14. Dystrophin glycoprotein complex-associated Gbetagamma subunits activate phosphatidylinositol-3-kinase/Akt signaling in skeletal muscle in a laminin-dependent manner.

    PubMed

    Xiong, Yongmin; Zhou, Yanwen; Jarrett, Harry W

    2009-05-01

    Previously, we showed that laminin-binding to the dystrophin glycoprotein complex (DGC) of skeletal muscle causes a heterotrimeric G-protein (Galphabetagamma) to bind, changing the activation state of the Gsalpha subunit. Others have shown that laminin-binding to the DGC also leads to Akt activation. Gbetagamma, released when Gsalpha is activated, is known to bind phosphatidylinositol-3-kinase (PI3K), which activates Akt in other cells. Here, we investigate whether muscle Akt activation results from Gbetagamma, using immunoprecipitation and immunoblotting, and purified Gbetagamma. In the presence of laminin, PI3K-binding to the DGC increases and Akt becomes phosphorylated and activated (pAkt), and glycogen synthase kinase is phosphorylated. Antibodies, which specifically block laminin-binding to alpha-dystroglycan, prevent PI3K-binding to the DGC. Purified bovine brain Gbetagamma also caused PI3K and Akt activation. These results show that DGC-Gbetagamma is binding PI3K and activating pAkt in a laminin-dependent manner. Mdx mice, which have greatly diminished amounts of DGC proteins, display elevated pAkt signaling and increased expression of integrin beta1 compared to normal muscle. This integrin binds laminin, Gbetagamma, and PI3K. Collectively, these suggest that PI3K is an important target for the Gbetagamma, which normally binds to DGC syntrophin, and activates PI3K/Akt signaling. Disruption of the DGC in mdx mouse is causing dis-regulation of the laminin-DGC-Gbetagamma-PI3K-Akt signaling and is likely to be important to the pathogenesis of muscular dystrophy. Upregulating integrin beta1 expression and activating the PI3K/Akt pathway in muscular dystrophy may partially compensate for the loss of the DGC. The results suggest new therapeutic approaches to muscle disease.

  15. Structure-Based Design of an Organoruthenium Phosphatidyl-inositol-3-Kinase Inhibitor Reveals a Switch Governing Lipid Kinase Potency and Selectivity

    SciTech Connect

    Xie,P.; Williams, D.; Atilla-Gokcumen, G.; Milk, L.; Xiao, M.; Smalley, K.; Herlyn, M.; Meggers, E.; Marmorstein, R.

    2008-01-01

    Mutations that constitutively activate the phosphatidyl-inositol-3-kinase (PI3K) signaling pathway, including alterations in PI3K, PTEN, and AKT, are found in a variety of human cancers, implicating the PI3K lipid kinase as an attractive target for the development of therapeutic agents to treat cancer and other related diseases. In this study, we report on the combination of a novel organometallic kinase inhibitor scaffold with structure-based design to develop a PI3K inhibitor, called E5E2, with an IC50 potency in the mid-low-nanomolar range and selectivity against a panel of protein kinases. We also show that E5E2 inhibits phospho-AKT in human melanoma cells and leads to growth inhibition. Consistent with a role for the PI3K pathway in tumor cell invasion, E5E2 treatment also inhibits the migration of melanoma cells in a 3D spheroid assay. The structure of the PI3K?/E5E2 complex reveals the molecular features that give rise to this potency and selectivity toward lipid kinases with implications for the design of a subsequent generation of PI3K-isoform-specific organometallic inhibitors.

  16. Restraining PI3K: mTOR signalling goes back to the membrane.

    PubMed

    Harrington, Laura S; Findlay, Greg M; Lamb, Richard F

    2005-01-01

    The lipid kinase phosphoinositide 3-kinase (PI3K) is activated in response to various extracellular signals including peptide growth factors such as insulin and insulin-like growth factors (IGFs). Phosphatidylinositol (3,4,5)-trisphosphate [PtdIns(3,4,5)P(3)] generated by PI3K is central to the diverse responses elicited by insulin, including glucose homeostasis, proliferation, survival and cell growth. The actions of lipid phosphatases have been considered to be the main means of attenuating PI3K signalling, whereby the principal 3-phosphatase - phosphatase and tensin homologue deleted on chromosome 10 (PTEN) - dephosphorylates PtdIns(3,4,5)P(3), reversing the action of PI3K. Recently, however, another pathway of regulation of PI3K has been identified in which activation of PI3K itself is prevented. This finding, together with earlier work, strongly suggests that a major form of negative feedback inhibition of PI3K results from activated growth signalling via mammalian target of rapamycin (mTOR) and the p70 S6 kinase (S6K) - a pathway that could have consequences for the development of type 2 diabetes and tuberous sclerosis complex.

  17. Abrogating endocrine resistance by targeting ERα and PI3K in breast cancer

    PubMed Central

    Fox, Emily M.; Arteaga, Carlos L.; Miller, Todd W.

    2012-01-01

    Antiestrogen therapies targeting estrogen receptor α (ER) signaling are a mainstay for patients with ER+ breast cancer. While many cancers exhibit resistance to antiestrogen therapies, a large body of clinical and experimental evidence indicates that hyperactivation of the phosphatidylinositol 3-kinase (PI3K) pathway promotes antiestrogen resistance. In addition, continued ligand-independent ER signaling in the setting of estrogen deprivation may contribute to resistance to endocrine therapy. PI3K activates several proteins which promote cell cycle progression and survival. In ER+ breast cancer cells, PI3K promotes ligand-dependent and -independent ER transcriptional activity. Models of antiestrogen-resistant breast cancer often remain sensitive to estrogen stimulation and PI3K inhibition, suggesting that clinical trials with combinations of drugs targeting both the PI3K and ER pathways are warranted. Herein, we review recent findings on the roles of PI3K and ER in antiestrogen resistance, and clinical trials testing drug combinations which target both pathways. We also discuss the need for clinical investigation of ER downregulators in combination with PI3K inhibitors. PMID:23087906

  18. Will PI3K pathway inhibitors be effective as single agents in patients with cancer?

    PubMed Central

    Garrett, Joan T.; Chakrabarty, Anindita; Arteaga, Carlos L.

    2011-01-01

    The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) axis regulates essential cellular functions including cell survival, proliferation, metabolism, migration, and angiogenesis. The PI3K pathway is activated in human cancers by mutation, amplification, and deletion of genes encoding components of this pathway. The critical role of PI3K in cancer has led to the development of drugs targeting the effector mechanisms of this signaling network. Recent studies have shown that inhibition at multiple levels of the PI3K pathway results in FOXO-dependent feedback reactivation of several receptor tyrosine kinases (RTKs) which, in turn, limit the sustained inhibition of this pathway and attenuates the action of therapeutic antagonists. This suggests that if used as single agents, PI3K pathway inhibitors may have limited clinical activity. We propose herein that to successfully target the output of the PI3K pathway in cancer cells, combination therapies that hinder these compensatory mechanisms should be used. Thus, combination therapies that target RTKs, PI3K, and mTOR activities may be required to maximize the clinical benefit derived from treatment with these inhibitors. PMID:22248929

  19. Airway PI3K Pathway Activation Is an Early and Reversible Event in Lung Cancer Development

    PubMed Central

    Gustafson, Adam M.; Soldi, Raffaella; Anderlind, Christina; Scholand, Mary Beth; Qian, Jun; Zhang, Xiaohui; Cooper, Kendal; Walker, Darren; McWilliams, Annette; Liu, Gang; Szabo, Eva; Brody, Jerome; Massion, Pierre P.; Lenburg, Marc E.; Lam, Stephen; Bild, Andrea H.; Spira, Avrum

    2013-01-01

    Although only a subset of smokers develop lung cancer, we cannot determine which smokers are at highest risk for cancer development, nor do we know the signaling pathways altered early in the process of tumorigenesis in these individuals. On the basis of the concept that cigarette smoke creates a molecular field of injury throughout the respiratory tract, this study explores oncogenic pathway deregulation in cytologically normal proximal airway epithelial cells of smokers at risk for lung cancer. We observed a significant increase in a genomic signature of phosphatidylinositol 3-kinase (PI3K) pathway activation in the cytologically normal bronchial airway of smokers with lung cancer and smokers with dysplastic lesions, suggesting that PI3K is activated in the proximal airway before tumorigenesis. Further, PI3K activity is decreased in the airway of high-risk smokers who had significant regression of dysplasia after treatment with the chemopreventive agent myo-inositol, and myo-inositol inhibits the PI3K pathway in vitro. These results suggest that deregulation of the PI3K pathway in the bronchial airway epithelium of smokers is an early, measurable, and reversible event in the development of lung cancer and that genomic profiling of these relatively accessible airway cells may enable personalized approaches to chemoprevention and therapy. Our work further suggests that additional lung cancer chemoprevention trials either targeting the PI3K pathway or measuring airway PI3K activation as an intermediate endpoint are warranted. PMID:20375364

  20. Modulation of glycolysis and lipogenesis by novel PI3K selective molecule represses tumor angiogenesis and decreases colorectal cancer growth.

    PubMed

    Hussain, Aashiq; Qazi, Asif Khurshid; Mupparapu, Nagaraju; Guru, Santosh Kumar; Kumar, Ashok; Sharma, Parduman Raj; Singh, Shashank Kumar; Singh, Paramjit; Dar, Mohd Jamal; Bharate, Sandip B; Zargar, Mohmmad Afzal; Ahmed, Qazi Naveed; Bhushan, Shashi; Vishwakarma, Ram A; Hamid, Abid

    2016-05-01

    Phosphatidylinositol 3-kinase (PI3K) pathway drives cancer progression through direct regulation of most oncogenic properties. Here, we report that PI3K pathway signaling up-regulates cancer cell proliferation, metastasis and angiogenesis through modulation of cancer metabolism. These oncogenic metabolic processes were disrupted, by a novel PI3K inhibitor, 3-Dihydro-2-(naphthalene-1-yl) quinazolin-4(1H)-one (DHNQ) in colon cancer cells. DHNQ inhibited the Warburg effect and lipid synthesis by reducing gene expression of glycolytic and lipogenesis regulatory enzymes. This downregulation at gene level by DHNQ inhibited metabolic flux to repress proliferation, migration and invasion characteristics of colon cancer. Furthermore, the metabolic attenuation caused repression of in vitro/in vivo angiogenesis providing new insights in PI3K regulated angiogenesis via metabolic alterations. Our results suggest that multifaceted targeting of oncogenic metabolism by their upstream PI3K regulatory signaling may be an effective cancer treatment approach.

  1. Chronic Blockade of Phosphatidylinositol 3-Kinase in the Nucleus Tractus Solitarii Is Prohypertensive in the Spontaneously Hypertensive Rat

    PubMed Central

    Zubcevic, Jasenka; Waki, Hidefumi; Diez-Freire, Carlos; Gampel, Alexandra; Raizada, Mohan K.; Paton, Julian F.R.

    2009-01-01

    Phosphatidylinositol 3-kinase (PI3K) within brain stem neurons has been implicated in hypertension in the spontaneously hypertensive rat (SHR). Previously, we demonstrated elevated expression of PI3K subunits in rostral ventrolateral medulla and paraventricular nucleus of SHRs compared with Wistar-Kyoto rats. Here, we considered expression levels of PI3K in the nucleus tractus solitarii, a pivotal region in reflex regulation of arterial pressure, and determined its functional role for arterial pressure homeostasis in SHRs and Wistar-Kyoto rats. We found elevated mRNA levels of p110β and p110δ catalytic PI3K subunits in the nucleus tractus solitarii of adult (12 to 14 weeks old) SHRs relative to the age-matched Wistar-Kyoto rats (fold differences relative to β-actin: 1.7±0.2 versus 1.01±0.08 for p110β, n=6, P<0.05; 1.62±0.15 versus 1.02±0.1 for p110δ, n=6, P<0.05). After chronic blockade of PI3K signaling in the nucleus tractus solitarii by lentiviral-mediated expression of a mutant form of p85α, systolic pressure increased from 175±3 mm Hg to 191±6 mm Hg (P<0.01) in SHRs but not in Wistar-Kyoto rats. In addition, heart rate increased (from 331±6 to 342±6 bpm; P<0.05) and spontaneous baroreflex gain decreased (from 0.7±0.07 to 0.5±0.04 ms/mm Hg; P<0.001) in the SHRs. Thus, PI3K signaling in the nucleus tractus solitarii of SHR restrains arterial pressure in this animal model of neurogenic hypertension. PMID:19015400

  2. Early Activation of Phosphatidylinositol 3-Kinase after Ischemic Stroke Reduces Infarct Volume and Improves Long-Term Behavior.

    PubMed

    Kim, Young Seo; Yoo, Arum; Son, Jeong Woo; Kim, Hyun Young; Lee, Young-Jun; Hwang, Sejin; Lee, Kyu-Yong; Lee, Young Joo; Ayata, Cenk; Kim, Hyung-Hwan; Koh, Seong-Ho

    2017-09-01

    Phosphatidylinositol 3-kinases (PI3Ks) have recently been implicated in apoptosis and ischemic cell death. We tested the efficacy of early intervention with a peptide PI3K activator in focal cerebral ischemia. After determining the most effective dose (24 μg/kg) and time window (2 h after MCAO) of treatment, a total of 48 rats were subjected to middle cerebral artery occlusion (MCAO). Diffusion weighted MRI (DWI) was performed 1 h after MCAO and rats with lesion sizes within a predetermined range were randomized to either PI3K activator or vehicle treatment arms. Fluid attenuated inversion recovery (FLAIR) MRI, neurological function, western blots, and immunohistochemistry were blindly assessed. Initial DWI lesion volumes were nearly identical between two groups prior to treatment. However, FLAIR showed significantly smaller infarct volumes in the PI3K activator group compared with vehicle (146 ± 81 mm(3) and 211 ± 96 mm(3), p = 0.045) at 48 h. The PI3K activator group also had better neurological function for up to 2 weeks. In addition, PI3K activator decreased the number of TUNEL-positive cells in the peri-infarct region compared with the control group. Western blot and immunohistochemistry showed increased expression of phosphorylated Akt (Ser473) and GSK-3β (Ser9) and decreased expression of cleaved caspase-9 and caspase-3. Our results suggest a neuroprotective role of early activation of PI3K in ischemic stroke. The use of DWI in the randomization of experimental groups may reduce bias.

  3. Phosphatidylinositol 3-kinase mediates activation of ATM by high NaCl and by ionizing radiation: Role in osmoprotective transcriptional regulation

    PubMed Central

    Irarrazabal, Carlos E.; Burg, Maurice B.; Ward, Stephen G.; Ferraris, Joan D.

    2006-01-01

    High NaCl causes DNA double-strand breaks and activates the transcription factor, TonEBP/OREBP, resulting in increased transcription of several protective genes, including those involved in accumulation of compatible organic osmolytes. Several kinases are known to contribute to signaling activation of TonEBP/OREBP, including ATM, which is a member of the phosphatidylinositol 3-kinase (PI3K)-like kinase family and is activated by DNA double-strand breaks. The purpose of the present studies was to investigate a possible role of PI3K Class IA (PI3K-IA). We found that high NaCl increases PI3K-IA lipid kinase activity. Inhibiting PI3K-IA either by expressing a dominant negative of its regulatory subunit, p85, or by small interfering RNA-mediated knockdown of its catalytic subunit, p110α, reduces high NaCl-induced increases in TonEBP/OREBP transcriptional activity and transactivation, but not nuclear translocation of TonEBP/OREBP, or increases in its abundance. Further, suppression of PI3K-IA inhibits the activation of ATM that is caused by either ionizing radiation or high NaCl. High NaCl-induced increase in TonEBP/OREBP activity is reduced equally by inhibition of ATM or PI3K-IA, and the effects are not additive. The conclusions are as follows: (i) PI3K-IA activity is necessary for both high NaCl- and ionizing radiation-induced activation of ATM and (ii) high NaCl activates PI3K-IA, which, in turn, contributes to full activation of TonEBP/OREBP via ATM. PMID:16728507

  4. Targeting the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin signaling network in cancer stem cells.

    PubMed

    Martelli, A M; Evangelisti, C; Follo, M Y; Ramazzotti, G; Fini, M; Giardino, R; Manzoli, L; McCubrey, J A; Cocco, L

    2011-01-01

    Cancer stem cells (CSCs) comprise a subset of hierarchically organized, rare cancer cells with the ability to initiate cancer in xenografts of genetically modified murine models. CSCs are thought to be responsible for tumor onset, self-renewal/maintenance, mutation accumulation, and metastasis. The existence of CSCs could explain the high frequency of neoplasia relapse and resistance to all of currently available therapies, including chemotherapy. The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway is a key regulator of physiological cell processes which include proliferation, differentiation, apoptosis, motility, metabolism, and autophagy. Nevertheless, aberrantly upregulated PI3K/Akt/mTOR signaling characterizes many types of cancers where it negatively influences prognosis. Several lines of evidence indicate that this signaling system plays a key role also in CSC biology. Of note, CSCs are more sensitive to pathway inhibition with small molecules when compared to healthy stem cells. This observation provides the proof-of-principle that functional differences in signaling transduction pathways between CSCs and healthy stem cells can be identified. Here, we review the evidence which links the signals deriving from the PI3K/Akt/mTOR network with CSC biology, both in hematological and solid tumors. We then highlight how therapeutic targeting of PI3K/Akt/mTOR signaling with small molecule inhibitors could improve cancer patient outcome, by eliminating CSCs.

  5. Glycogen Synthase Kinase 3 influences cell motility and chemotaxis by regulating PI3K membrane localization in Dictyostelium.

    PubMed

    Sun, Tong; Kim, Bohye; Kim, Lou W

    2013-10-01

    Glycogen Synthase Kinase 3 (GSK3) is a multifunctional kinase involved in diverse cellular activities such as metabolism, differentiation, and morphogenesis. Recent studies showed that GSK3 in Dictyostelium affects chemotaxis via TorC2 pathway and Daydreamer. Now we report that GSK3 affects PI3K membrane localization, of which the mechanism has remained to be fully understood in Dictyostelium. The membrane localization domain (LD) of Phosphatidylinositol-3-kinase 1 (PI3K1) is phosphorylated on serine residues in a GSK3 dependent mechanism and PI3K1-LD exhibited biased membrane localization in gsk3(-) cells compared to the wild type cells. Furthermore, multiple GSK3-phosphorylation consensus sites exist in PI3K1-LD, of which phosphomimetic substitutions restored cAMP induced transient membrane localization of PI3K1-LD in gsk3(-) cells. Serine to alanine substitution mutants of PI3K1-LD, in contrast, displayed constitutive membrane localization in wild type cells. Biochemical analysis revealed that GSK3 dependent serine phosphorylation of PI3K1-LD is constitutive during the course of cAMP stimulation. Together, these data suggest that GSK3 dependent serine phosphorylation is a prerequisite for chemoattractant cAMP induced PI3K membrane localization.

  6. Glycogen Synthase Kinase 3 influences cell motility and chemotaxis by regulating PI3K membrane localization in Dictyostelium

    PubMed Central

    Sun, Tong; Kim, Bohye; Kim, Lou W.

    2013-01-01

    Glycogen Synthase Kinase 3 (GSK3) is a multifunctional kinase involved in diverse cellular activities such as metabolism, differentiation, and morphogenesis. Recent studies showed that GSK3 in Dictyostelium affects chemotaxis via TorC2 pathway and Daydreamer. Now we report that GSK3 affects PI3K membrane localization, of which mechanism has remained to be fully understood in Dictyostelium. The membrane localization domain (LD) of Phosphatidylinositol-3-kinase 1 (PI3K1) is phosphorylated on serine residues in a GSK3 dependent mechanism and PI3K1-LD exhibited biased membrane localization in gsk3− cells compared to the wild type cells. Furthermore, multiple GSK3-phosphorylation consensus sites exist in PI3K1-LD, of which phosphomimetic substitutions restored cAMP induced transient membrane localization of PI3K1-LD in gsk3− cells. Serine to alanine substitution mutants of PI3K1-LD, in contrast, displayed constitutive membrane localization in wild type cells. Biochemical analysis revealed that GSK3 dependent serine phosphorylation of PI3K1-LD is constitutive during the course of cAMP stimulation. Together, these data suggest that GSK3 dependent serine phosphorylation is a prerequisite for chemoattractant cAMP induced PI3K membrane localization. PMID:24102085

  7. Phosphatidylinositol 3-kinase-δ inhibitor CAL-101 shows promising preclinical activity in chronic lymphocytic leukemia by antagonizing intrinsic and extrinsic cellular survival signals

    PubMed Central

    Herman, Sarah E. M.; Gordon, Amber L.; Wagner, Amy J.; Heerema, Nyla A.; Zhao, Weiqiang; Flynn, Joseph M.; Jones, Jeffrey; Andritsos, Leslie; Puri, Kamal D.; Lannutti, Brian J.; Giese, Neill A.; Zhang, Xiaoli; Wei, Lai; Byrd, John C.

    2010-01-01

    Targeted therapy with imatinib in chronic myeloid leukemia (CML) prompted a new treatment paradigm. Unlike CML, chronic lymphocytic leukemia (CLL) lacks an aberrant fusion protein kinase but instead displays increased phosphatidylinositol 3-kinase (PI3K) activity. To date, PI3K inhibitor development has been limited because of the requirement of this pathway for many essential cellular functions. Identification of the hematopoietic-selective isoform PI3K-δ unlocks a new therapeutic potential for B-cell malignancies. Herein, we demonstrate that PI3K has increased enzymatic activity and that PI3K-δ is expressed in CLL cells. A PI3K-δ selective inhibitor CAL-101 promoted apoptosis in primary CLL cells ex vivo in a dose- and time-dependent fashion that was independent of common prognostic markers. CAL-101–mediated cytotoxicity was caspase dependent and was not diminished by coculture on stromal cells. In addition, CAL-101 abrogated protection from spontaneous apoptosis induced by B cell–activating factors CD40L, TNF-α, and fibronectin. In contrast to malignant cells, CAL-101 does not promote apoptosis in normal T cells or natural killer cells, nor does it diminish antibody-dependent cellular cytotoxicity. However, CAL-101 did decrease activated T-cell production of various inflammatory and antiapoptotic cytokines. Collectively, these studies provide rationale for the clinical development of CAL-101 as a first-in-class targeted therapy for CLL and related B-cell lymphoproliferative disorders. PMID:20522708

  8. Class IA phosphatidylinositol 3-kinase in pancreatic β cells controls insulin secretion by multiple mechanisms.

    PubMed

    Kaneko, Kazuma; Ueki, Kohjiro; Takahashi, Noriko; Hashimoto, Shinji; Okamoto, Masayuki; Awazawa, Motoharu; Okazaki, Yukiko; Ohsugi, Mitsuru; Inabe, Kazunori; Umehara, Toshihiro; Yoshida, Masashi; Kakei, Masafumi; Kitamura, Tadahiro; Luo, Ji; Kulkarni, Rohit N; Kahn, C Ronald; Kasai, Haruo; Cantley, Lewis C; Kadowaki, Takashi

    2010-12-01

    Type 2 diabetes is characterized by insulin resistance and pancreatic β cell dysfunction, the latter possibly caused by a defect in insulin signaling in β cells. Inhibition of class IA phosphatidylinositol 3-kinase (PI3K), using a mouse model lacking the pik3r1 gene specifically in β cells and the pik3r2 gene systemically (βDKO mouse), results in glucose intolerance and reduced insulin secretion in response to glucose. β cells of βDKO mice had defective exocytosis machinery due to decreased expression of soluble N-ethylmaleimide attachment protein receptor (SNARE) complex proteins and loss of cell-cell synchronization in terms of Ca(2+) influx. These defects were normalized by expression of a constitutively active form of Akt in the islets of βDKO mice, preserving insulin secretion in response to glucose. The class IA PI3K pathway in β cells in vivo is important in the regulation of insulin secretion and may be a therapeutic target for type 2 diabetes.

  9. High fat diet induced obesity alters ovarian phosphatidylinositol-3 kinase signaling gene expression.

    PubMed

    Nteeba, J; Ross, J W; Perfield, J W; Keating, A F

    2013-12-01

    Insulin regulates ovarian phosphatidylinositol-3-kinase (PI3 K) signaling, important for primordial follicle viability and growth activation. This study investigated diet-induced obesity impacts on: (1) insulin receptor (Insr) and insulin receptor substrate 1 (Irs1); (2) PI3K components (Kit ligand (Kitlg), kit (c-Kit), protein kinase B alpha (Akt1) and forkhead transcription factor subfamily 3 (Foxo3a)); (3) xenobiotic biotransformation (microsomal epoxide hydrolase (Ephx1), Cytochrome P450 isoform 2E1 (Cyp2e1), Glutathione S-transferase (Gst) isoforms mu (Gstm) and pi (Gstp)) and (4) microRNA's 184, 205, 103 and 21 gene expression. INSR, GSTM and GSTP protein levels were also measured. Obese mouse ovaries had decreased Irs1, Foxo3a, Cyp2e1, MiR-103, and MiR-21 but increased Kitlg, Akt1, and miR-184 levels relative to lean littermates. These results support that diet-induced obesity potentially impairs ovarian function through aberrant gene expression. Copyright © 2013 Elsevier Inc. All rights reserved.

  10. High fat diet induced obesity alters ovarian phosphatidylinositol-3 kinase signaling gene expression

    PubMed Central

    Nteeba, J.; Ross, J.W.; Perfield, J.W.; Keating, A.F.

    2013-01-01

    Insulin regulates ovarian phosphatidylinositol-3-kinase (PI3K) signaling, important for primordial follicle viability and growth activation. This study investigated diet-induced obesity impacts on: 1) insulin receptor (Insr) and insulin receptor substrate 1 (Irs1); 2) PI3K components (Kit ligand (Kitlg), kit (c-Kit), protein kinase B alpha (Akt1) and forkhead transcription factor subfamily 3 (Foxo3a)); 3) xenobiotic biotransformation (microsomal epoxide hydrolase (Ephx1), Cytochrome P450 isoform 2E1 (Cyp2e1), Glutathione S-transferase (Gst) isoforms mu (Gstm) and pi (Gstp)) and 4) microRNA’s 184, 205, 103 and 21 gene expression. INSR, GSTM and GSTP protein levels were also measured. Obese mouse ovaries had decreased Irs1, Foxo3a, Cyp2e1, MiR-103, and MiR-21 but increased Kitlg, Akt1, and miR-184 levels relative to lean littermates. These results support that diet-induced obesity potentially impairs ovarian function through aberrant gene expression. PMID:23954404

  11. Creatine inhibits adipogenesis by downregulating insulin-induced activation of the phosphatidylinositol 3-kinase signaling pathway.

    PubMed

    Lee, Nayeon; Kim, Inhee; Park, Soojeong; Han, Dasol; Ha, Soobong; Kwon, Mookwang; Kim, Juwan; Byun, Sung-Hyun; Oh, Wonil; Jeon, Hong Bae; Kweon, Dae-Hyuk; Cho, Jae Youl; Yoon, Keejung

    2015-04-15

    Creatine is a nitrogenous organic acid known to function in adenosine triphosphate (ATP) metabolism. Recent evidence indicates that creatine regulates the differentiation of mesenchymal stem cells (MSCs) in processes such as osteogenesis and myogenesis. In this study, we show that creatine also has a negative regulatory effect on fat cell formation. Creatine inhibits the accumulation of cytoplasmic triglycerides in a dose-dependent manner irrespective of the adipogenic cell models used, including a C3H10T1/2 MSC line, 3T3-L1 preadipocytes, and primary human MSCs. Consistently, a dramatic reduction in mRNA expression of adipogenic transcription factors, peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα), glucose transporters, 1 and 4 (Glut1, Glut4), and adipocyte markers, aP2 and adipsin, was observed in the presence of creatine. Creatine appears to exert its inhibitory effects on adipogenesis during early differentiation, but not late differentiation, or proliferation stages through inhibition of the PI3K-Akt-PPARγ signaling pathway. In an in vivo model, administration of creatine into mice resulted in body mass increase without fat accumulation. In summary, our results indicate that creatine downregulates adipogenesis through inhibition of phosphatidylinositol 3-kinase (PI3K) activation and imply the potent therapeutic value of creatine in treating obesity and obesity-related metabolic disorders.

  12. Dual inhibition of class IA phosphatidylinositol 3-kinase and mammalian target of rapamycin as a new therapeutic option for T-cell acute lymphoblastic leukemia.

    PubMed

    Chiarini, Francesca; Falà, Federica; Tazzari, Pier Luigi; Ricci, Francesca; Astolfi, Annalisa; Pession, Andrea; Pagliaro, Pasqualepaolo; McCubrey, James A; Martelli, Alberto M

    2009-04-15

    Recent investigations have documented that constitutively activated phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling is a common feature of T-cell acute lymphoblastic leukemia (T-ALL), where it strongly influences growth and survival. These findings lend compelling weight for the application of PI3K/Akt/mTOR inhibitors in T-ALL. However, our knowledge of PI3K/Akt/mTOR signaling in T-ALL is limited and it is not clear whether it could be an effective target for innovative therapeutic strategies. Here, we have analyzed the therapeutic potential of the dual PI3K/mTOR inhibitor PI-103, a small synthetic molecule of the pyridofuropyrimidine class, on both T-ALL cell lines and patient samples, which displayed constitutive activation of PI3K/Akt/mTOR signaling. PI-103 inhibited the growth of T-ALL cells, including 170-kDa P-glycoprotein overexpressing cells. PI-103 cytotoxicity was independent of p53 gene status. PI-103 was more potent than inhibitors that are selective only for PI3K (Wortmannin, LY294002) or for mTOR (rapamycin). PI-103 induced G(0)-G(1) phase cell cycle arrest and apoptosis, which was characterized by activation of caspase-3 and caspase-9. PI-103 caused Akt dephosphorylation, accompanied by dephosphorylation of the Akt downstream target, glycogen synthase kinase-3beta. Also, mTOR downstream targets were dephosphorylated in response to PI-103, including p70S6 kinase, ribosomal S6 protein, and 4E-BP1. PI-103 strongly synergized with vincristine. These findings indicate that multitargeted therapy toward PI3K and mTOR alone or with existing drugs may serve as an efficient treatment toward T-ALL cells, which require up-regulation of PI3K/Akt/mTOR signaling for their survival and growth.

  13. Phosphatidylinositol-3-kinase pathway aberrations in gastric and colorectal cancer: meta-analysis, co-occurrence and ethnic variation.

    PubMed

    Chong, Mei-Ling; Loh, Marie; Thakkar, Bhavin; Pang, Brendan; Iacopetta, Barry; Soong, Richie

    2014-03-01

    Inhibition of the phosphatidylinositol-3-kinase (PI3K) signaling pathway is a cancer treatment strategy that has entered into clinical trials. We performed a meta-analysis on the frequency of prominent genetic (PIK3CA mutation, PIK3CA amplification and PTEN deletion) and protein expression (high PI3K, PTEN loss and high pAkt) aberrations in the PI3K pathway in gastric cancer (GC) and colorectal cancer (CRC). We also performed laboratory analysis to investigate the co-occurrence of these aberrations. The meta-analysis indicated that East Asian and Caucasian GC patients differ significantly for the frequencies of PIK3CA Exon 9 and 20 mutations (7% vs. 15%, respectively), PTEN deletion (21% vs. 4%) and PTEN loss (47% vs. 78%), while CRC patients differed for PTEN loss (57% vs. 26%). High study heterogeneity (I(2) > 80) was observed for all aberrations except PIK3CA mutations. Laboratory analysis of tumors from East Asian patients revealed significant differences between GC (n = 79) and CRC (n = 116) for the frequencies of PIK3CA amplification (46% vs. 4%) and PTEN loss (54% vs. 78%). The incidence of GC cases with 0, 1, 2 and 3 concurrent aberrations was 14%, 52%, 27% and 8%, respectively, while for CRC it was 10%, 60%, 25% and 4%, respectively. Our study consolidates knowledge on the frequency, co-occurrence and clinical relevance of PI3K pathway aberrations in GC and CRC. Up to 86% of GC and 90% of CRC have at least one aberration in the PI3K pathway, and there are significant differences in the frequencies of these aberrations according to cancer type and ethnicity.

  14. A Potent Inhibitor of Phosphoinositide 3-Kinase (PI3K) and Mitogen Activated Protein (MAP) Kinase Signalling, Quercetin (3, 3', 4', 5, 7-Pentahydroxyflavone) Promotes Cell Death in Ultraviolet (UV)-B-Irradiated B16F10 Melanoma Cells

    PubMed Central

    Rafiq, Rather A.; Quadri, Afnan; Nazir, Lone A.; Peerzada, Kaiser; Ganai, Bashir A.; Tasduq, Sheikh A.

    2015-01-01

    Ultraviolet (UV) radiation–induced skin damage contributes strongly to the formation of melanoma, a highly lethal form of skin cancer. Quercetin (Qu), the most widely consumed dietary bioflavonoid and well known inhibitor of phosphoinositide 3-kinase (PI3K) and mitogen activated protein (MAP) kinase signalling, has been reported to be chemopreventive in several forms of non-melanoma skin cancers. Here, we report that the treatment of ultraviolet (UV)-B-irradiated B16F10 melanoma cells with quercetin resulted in a dose dependent reduction in cell viability and increased apoptosis. The present study has brought out that the pro-apoptotic effects of quercetin in UVB-irradiated B16F10 cells are mediated through the elevation of intracellular reactive oxygen species (ROS) formation, calcium homeostasis imbalance, modulation of anti-oxidant defence response and depolarization of mitochondrial membrane potential (ΔΨM). Promotion of UVB-induced cell death by quercetin was further revealed by cleavage of chromosomal DNA, caspase activation, poly (ADP) ribose polymerase (PARP) cleavage, and an increase in sub-G1 cells. Quercetin markedly attenuated MEK-ERK signalling, influenced PI3K/Akt pathway, and potentially enhanced the UVB-induced NF-κB nuclear translocation. Furthermore, combined UVB and quercetin treatment decreased the ratio of Bcl-2 to that of Bax, and upregulated the expression of Bim and apoptosis inducing factor (AIF). Overall, these results suggest the possibility of using quercetin in combination with UVB as a possible treatment option for melanoma in future. PMID:26148186

  15. The Role of PI3K/Akt in Human Herpesvirus Infection: from the Bench to the Bedside

    PubMed Central

    Liu, XueQiao; Cohen, Jeffrey I.

    2015-01-01

    The phosphatidylinositol-3-kinase (PI3K)-Akt signaling pathway regulates several key cellular functions including protein synthesis, cell growth, glucose metabolism, and inflammation. Many viruses have evolved mechanisms to manipulate this signaling pathway to ensure successful virus replication. The human herpesviruses undergo both latent and lytic infection, but differ in cell tropism, growth kinetics, and disease manifestations. Herpesviruses express multiple proteins that target the PI3K/Akt cell signaling pathway during the course of their life cycle to facilitate viral infection, replication, latency, and reactivation. Rare human genetic disorders with mutations in either the catalytic or regulatory subunit of PI3K that result in constitutive activation of the protein predispose to severe herpesvirus infections as well as to virus-associated malignancies. Inhibiting the PI3K/Akt pathway or its downstream proteins using drugs already approved for other diseases can block herpesvirus lytic infection and may reduce malignancies associated with latent herpesvirus infections. PMID:25798530

  16. Coordinate activation of Shh and PI3K signaling in PTEN-deficient glioblastoma: new therapeutic opportunities.

    PubMed

    Filbin, Mariella Gruber; Dabral, Sukriti K; Pazyra-Murphy, Maria F; Ramkissoon, Shakti; Kung, Andrew L; Pak, Ekaterina; Chung, Jarom; Theisen, Matthew A; Sun, Yanping; Franchetti, Yoko; Sun, Yu; Shulman, David S; Redjal, Navid; Tabak, Barbara; Beroukhim, Rameen; Wang, Qi; Zhao, Jean; Dorsch, Marion; Buonamici, Silvia; Ligon, Keith L; Kelleher, Joseph F; Segal, Rosalind A

    2013-11-01

    In glioblastoma, phosphatidylinositol 3-kinase (PI3K) signaling is frequently activated by loss of the tumor suppressor phosphatase and tensin homolog (PTEN). However, it is not known whether inhibiting PI3K represents a selective and effective approach for treatment. We interrogated large databases and found that sonic hedgehog (SHH) signaling is activated in PTEN-deficient glioblastoma. We demonstrate that the SHH and PI3K pathways synergize to promote tumor growth and viability in human PTEN-deficient glioblastomas. A combination of PI3K and SHH signaling inhibitors not only suppressed the activation of both pathways but also abrogated S6 kinase (S6K) signaling. Accordingly, targeting both pathways simultaneously resulted in mitotic catastrophe and tumor apoptosis and markedly reduced the growth of PTEN-deficient glioblastomas in vitro and in vivo. The drugs tested here appear to be safe in humans; therefore, this combination may provide a new targeted treatment for glioblastoma.

  17. Pooled Analysis of Phosphatidylinositol 3-kinase Pathway Variants and Risk of Prostate Cancer

    PubMed Central

    Koutros, Stella; Schumacher, Fredrick R.; Hayes, Richard B.; Ma, Jing; Huang, Wen-Yi; Albanes, Demetrius; Canzian, Federico; Chanock, Stephen J.; Crawford, E. David; Diver, W. Ryan; Feigelson, Heather Spencer; Giovanucci, Edward; Haiman, Christopher A.; Henderson, Brian E.; Hunter, David J.; Kaaks, Rudolf; Kolonel, Laurence N.; Kraft, Peter; Le Marchand, Loïc; Riboli, Elio; Siddiq, Afshan; Stampfer, Mier J.; Stram, Daniel O.; Thomas, Gilles; Travis, Ruth C.; Thun, Michael J.; Yeager, Meredith; Berndt, Sonja I.

    2010-01-01

    The phosphatidylinositol 3-kinase (PI3K) pathway regulates various cellular processes, including cellular proliferation and intracellular trafficking and may impact prostate carcinogenesis. Thus, we explored the association between single nucleotide polymorphisms (SNPs) in PI3K genes and prostate cancer. Pooled data from the National Cancer Institute Breast and Prostate Cancer Cohort Consortium were examined for associations between 89 SNPs in PI3K genes (PIK3C2B, PIK3AP1, PIK3C2A, PIK3CD, and PIK3R3) and prostate cancer risk in 8,309 cases and 9,286 controls. Odds ratios (OR) and 95% confidence intervals (CI) were estimated using logistic regression. SNP rs7556371 in PIK3C2B was significantly associated with prostate cancer risk (ORper allele=1.08 (95% CI: 1.03, 1.14), p-trend = 0.0017) after adjustment for multiple testing (Padj=0.024). Simultaneous adjustment of rs7556371 for nearby SNPs strengthened the association (ORper allele=1.21 (95% CI: 1.09, 1.34); p-trend =0.0003). The adjusted association was stronger for men who were diagnosed before 65 years (ORper allele= 1.47 (95% CI: 1.20, 1.79), p-trend = 0.0001) or had a family history (ORper allele= 1.57 (95% CI: 1.11, 2.23), p-trend = 0.0114), and was strongest in those with both characteristics (ORper allele= 2.31 (95% CI: 1.07, 5.07), p-interaction = 0.005). Increased risks were observed among men in the top tertile of circulating insulin like growth factor-1 (IGF-1) levels (ORper allele= 1.46 (95% CI: 1.04, 2.06), p-trend=0.075). No differences were observed with disease aggressiveness (≥8/stage T3/T4/fatal). In conclusion, we observed a significant association between PIK3C2B and prostate cancer risk, especially for familial, early onset disease, which may be attributable to IGF-dependent PI3K signaling. PMID:20197460

  18. Effects of inhibiting PI3K-Akt-mTOR pathway on lipid metabolism homeostasis in goose primary hepatocytes.

    PubMed

    Liu, D D; Han, C C; Wan, H F; He, F; Xu, H Y; Wei, S H; Du, X H; Xu, F

    2016-08-01

    Phosphatidylinositol-3 kinases (PI3K)-Protein kinase B (Akt)-mammalian target of rapamycin (mTOR) pathway plays an important role in the synthesis and secretion of triacylglycerol. However, the mechanism of PI3K-Akt-mTOR pathway in regulating lipid metabolism of goose liver was poorly understood. The purpose of this study was to determine how PI3K-Akt-mTOR pathway regulating lipid metabolic homeostasis in goose hepatocytes. Goose primary hepatocytes were treated with different PI3K-Akt-mTOR signal inhibitors (LY294002, rapamycin and NVP-BEZ235) for 24 h. The results showed that these inhibitors evidently inhibited PI3K-Akt-mTOR downstream signaling. Meanwhile, these PI3K-Akt-mTOR inhibitors reduced intracellular lipid accumulation, decreased the mRNA expression and protein content of genes involved in the de novo fatty acid synthesis, while increased the transcriptional and protein level of key factors involved in fatty acid oxidation and very low density lipoprotein (VLDL) assembly and secretion. These findings suggested that the reduction of lipids accumulation induced-by inhibiting PI3K-Akt-mTOR pathway was closely linked to the decrease of lipogenesis, the increase of fatty acids oxidation, and the increase of VLDL assembly and secretion in goose hepatocytes.

  19. Loss of Cbl-PI3K interaction modulates the periosteal response to fracture by enhancing osteogenic commitment and differentiation.

    PubMed

    Scanlon, Vanessa; Walia, Bhavita; Yu, Jungeun; Hansen, Marc; Drissi, Hicham; Maye, Peter; Sanjay, Archana

    2017-02-01

    The periosteum contains multipotent skeletal progenitors that contribute to bone repair. The signaling pathways regulating the response of periosteal cells to fracture are largely unknown. Phosphatidylinositol-3 Kinase (PI3K), a prominent lipid kinase, is a major signaling protein downstream of several factors that regulate osteoblast differentiation. Cbl is an E3 ubiquitin ligase and a major adaptor protein that binds to the p85 regulatory subunit and modulates PI3K activity. Substitution of tyrosine 737 to phenylalanine (Y737F) in Cbl abolishes the interaction between Cbl and p85 subunit without affecting the Cbl's ubiquitin ligase function. Here, we investigated the role of PI3K signaling during the very early stages of fracture healing using Osterix(RFP) reporter mice. We found that the absence of PI3K regulation by Cbl resulted in robust periosteal thickening, with increased proliferation of periosteal cells. While the multipotent properties of periosteal progenitors to differentiate into chondrocytes and adipocytes did not change, osteogenic differentiation in the absence of Cbl-PI3K interaction was highly augmented. The increased stability and nuclear localization of Osterix observed in periosteal cells lacking Cbl-PI3K interaction may explain this enhanced osteogenic differentiation since the expression of Osterix transcriptional target genes including osteocalcin and BSP are increased in YF cells. Overall, our findings highlight a hitherto unexplored and novel role for Cbl and PI3K in modulating the osteogenic response of periosteal cells during the early stages of fracture repair.

  20. Central dopamine action modulates neuropeptide-controlled appetite via the hypothalamic PI3K/NF-κB-dependent mechanism.

    PubMed

    Hsieh, Y-S; Chen, P-N; Yu, C-H; Kuo, D-Y

    2014-11-01

    Hypothalamic neuropeptides, including neuropeptide Y (NPY) and proopiomelanocortin (POMC), have been found to control the appetite-suppressing effect of amphetamine (AMPH). In this study, we have examined whether dopamine receptor (DAR), phosphatidylinositol 3-kinase (PI3K) and nuclear factor-kappaB (NF-κB) are involved in AMPH's action. We administered AMPH to rats once a day for 4 days and assessed and compared changes in hypothalamic NPY, melanocortin receptor 4 (MC4R), PI3K, pAkt and NF-κB expression. We found that the inhibition of DAR increased NPY, but decreased MC4R, PI3K and NF-κB expression, compared with AMPH-treated rats. Moreover, MC4R, PI3K, pAkt and NF-κB increased with the maximum response on Day 2, which was consistent with the response of feeding behavior, but was opposite to the expression of NPY. Furthermore, we found that the intracerebroventricular infusion of the PI3K inhibitor or NF-κB antisense could attenuate AMPH-induced anorexia, and partially reverse the expression of NPY, MC4R, PI3K, Akt and NF-κB back toward a normal level. We, therefore, suggest that DAR-PI3K-NF-κB signaling in the hypothalamus plays functional roles in the modulation of NPY and POMC neurotransmissions and in the control of AMPH-evoked appetite suppression.

  1. PI3K-C2γ is a Rab5 effector selectively controlling endosomal Akt2 activation downstream of insulin signalling

    PubMed Central

    Braccini, Laura; Ciraolo, Elisa; Campa, Carlo C.; Perino, Alessia; Longo, Dario L.; Tibolla, Gianpaolo; Pregnolato, Marco; Cao, Yanyan; Tassone, Beatrice; Damilano, Federico; Laffargue, Muriel; Calautti, Enzo; Falasca, Marco; Norata, Giuseppe D.; Backer, Jonathan M.; Hirsch, Emilio

    2015-01-01

    In the liver, insulin-mediated activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway is at the core of metabolic control. Multiple PI3K and Akt isoenzymes are found in hepatocytes and whether isoform-selective interplays exist is currently unclear. Here we report that insulin signalling triggers the association of the liver-specific class II PI3K isoform γ (PI3K-C2γ) with Rab5-GTP, and its recruitment to Rab5-positive early endosomes. In these vesicles, PI3K-C2γ produces a phosphatidylinositol-3,4-bisphosphate pool specifically required for delayed and sustained endosomal Akt2 stimulation. Accordingly, loss of PI3K-C2γ does not affect insulin-dependent Akt1 activation as well as S6K and FoxO1-3 phosphorylation, but selectively reduces Akt2 activation, which specifically inhibits glycogen synthase activity. As a consequence, PI3K-C2γ-deficient mice display severely reduced liver accumulation of glycogen and develop hyperlipidemia, adiposity as well as insulin resistance with age or after consumption of a high-fat diet. Our data indicate PI3K-C2γ supports an isoenzyme-specific forking of insulin-mediated signal transduction to an endosomal pool of Akt2, required for glucose homeostasis. PMID:26100075

  2. Frequent PTEN genomic alterations and activated phosphatidylinositol 3-kinase pathway in basal-like breast cancer cells

    PubMed Central

    Marty, Bérengère; Maire, Virginie; Gravier, Eléonore; Rigaill, Guillem; Vincent-Salomon, Anne; Kappler, Marion; Lebigot, Ingrid; Djelti, Fathia; Tourdès, Audrey; Gestraud, Pierre; Hupé, Philippe; Barillot, Emmanuel; Cruzalegui, Francisco; Tucker, Gordon C; Stern, Marc-Henri; Thiery, Jean-Paul; Hickman, John A; Dubois, Thierry

    2008-01-01

    Introduction Basal-like carcinomas (BLCs) and human epidermal growth factor receptor 2 overexpressing (HER2+) carcinomas are the subgroups of breast cancers that have the most aggressive clinical behaviour. In contrast to HER2+ carcinomas, no targeted therapy is currently available for the treatment of patients with BLCs. In order to discover potential therapeutic targets, we aimed to discover deregulated signalling pathways in human BLCs. Methods In this study, we focused on the oncogenic phosphatidylinositol 3-kinase (PI3K) pathway in 13 BLCs, and compared it with a control series of 11 hormonal receptor negative- and grade III-matched HER2+ carcinomas. The two tumour populations were first characterised by immunohistochemistry and gene expression. The PI3K pathway was then investigated by gene copy-number analysis, gene expression profiling and at a proteomic level using reverse-phase protein array technology and tissue microarray. The effects of the PI3K inhibition pathway on proliferation and apoptosis was further analysed in three human basal-like cell lines. Results The PI3K pathway was found to be activated in BLCs and up-regulated compared with HER2+ tumours as shown by a significantly increased activation of the downstream targets Akt and mTOR (mammalian target of rapamycin). BLCs expressed significantly lower levels of the tumour suppressor PTEN and PTEN levels were significantly negatively correlated with Akt activity within that population. PTEN protein expression correlated significantly with PTEN DNA copy number and more importantly, reduced PTEN DNA copy numbers were observed specifically in BLCs. Similar to human samples, basal-like cell lines exhibited an activation of PI3K/Akt pathway and low/lack PTEN expression. Both PI3K and mTOR inhibitors led to basal-like cell growth arrest. However, apoptosis was specifically observed after PI3K inhibition. Conclusions These data provide insight into the molecular pathogenesis of BLCs and implicate the

  3. Inhibitory effect of capsaicin on B16-F10 melanoma cell migration via the phosphatidylinositol 3-kinase/Akt/Rac1 signal pathway

    PubMed Central

    Shin, Dong-Hoon; Kim, Ok-Hee; Jun, Hye-Seung

    2008-01-01

    Capsaicin (trans-8-methyl-N-vanillyl-6-nonenamide), the major pungent ingredient of red pepper, has been reported to possess anti-carcinogenic and anti-mutagenic activities. In this study, the anti-migration activity of capsaicin on highly metastatic B16-F10 melanoma cells was investigated. Capsaicin significantly inhibited the migration of melanoma cells without showing obvious cellular cytotoxicity at low doses. This effect correlated with the down-regulation of phosphatidylinositol 3-kinase (PI3-K) and its downstream target, Akt. Although B16-F10 cell migration was increased by the PI3-K activator through the activation of Akt, these PI3-K activator-induced phenomena were attenuated by capsaicin. Moreover, capsaicin was found to significantly inhibit Rac1 activity in a pull-down assay. These results demonstrate that capsaicin inhibits the migration of B16-F10 cells through the inhibition of the PI3-K/Akt/Rac1 signal pathway. The present investigation suggests that capsaicin targets PI3-K/Akt/Rac1-mediated cellular events in B16-F10 melanoma cells. Consequently, capsaicin administration should be considered an effective approach for the suppression of invasion and metastasis in malignant melanoma chemotherapy. PMID:18985006

  4. ARIA/HRG regulates AChR epsilon subunit gene expression at the neuromuscular synapse via activation of phosphatidylinositol 3-kinase and Ras/MAPK pathway

    PubMed Central

    1996-01-01

    AChR-inducing activity (ARIA)/heregulin, a ligand for erbB receptor tyrosine kinases (RTKs), is likely to be one nerve-supplied signal that induces expression of acetylcholine receptor (AChR) genes at the developing neuromuscular junction. Since some RTKs act through Ras and phosphatidylinositol 3-kinase (PI3K), we investigated the role of these pathways in ARIA signaling. Expression of activated Ras or Raf mimicked ARIA-induction of AChR epsilon subunit genes in muscle cells; whereas dominant negative Ras or Raf blocked the effect of ARIA. ARIA rapidly activated erk1 and erk2 and inhibition of both erks also abolished the effect of ARIA. ARIA stimulated association of PI3K with erbB3, expression of an activated PI3K led to ARIA-independent AChR epsilon subunit expression, and inhibition of PI3K abolished the action of ARIA. Thus, synaptic induction of AChR genes requires activation of both Ras/MAPK and PI3K signal transduction pathways. PMID:8707830

  5. The Role of PI3K/Akt/mTOR Signaling in Gastric Carcinoma

    PubMed Central

    Matsuoka, Tasuku; Yashiro, Masakazu

    2014-01-01

    The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is one of the key signaling pathways induced by various receptor-tyrosine kinases. Accumulating evidence shows that this pathway is an important promoter of cell growth, metabolism, survival, metastasis, and resistance to chemotherapy. Genetic alterations in the PI3K/Akt/mTOR pathway in gastric carcinoma have often been demonstrated. Many kinds of molecular targeting therapies are currently undergoing clinical testing in patients with solid tumors. However, with the exception of the ErbB2-targeting antibody, targeting agents, including PI3K/Akt/mTOR inhibitors, have not been approved for treatment of patients with gastric carcinoma. This review summarizes the current knowledge on PI3K/Akt/mTOR signaling in the pathogenesis of gastric carcinoma and the possible therapeutic targets for gastric carcinoma. Improved knowledge of the PI3K/Akt/mTOR pathway in gastric carcinoma will be useful in understanding the mechanisms of tumor development and for identifying ideal targets of anticancer therapy for gastric carcinoma. PMID:25003395

  6. Angelica Sinensis promotes myotube hypertrophy through the PI3K/Akt/mTOR pathway

    PubMed Central

    2014-01-01

    Background Angelica Sinensis (AS), a folk medicine, has long been used in ergogenic aids for athletes, but there is little scientific evidence supporting its effects. We investigated whether AS induces hypertrophy in myotubes through the phosphatidylinositol 3-kinase (PI3K)/Akt (also termed PKB)/mammalian target of the rapamycin (mTOR) pathway. Methods An in vitro experiment investigating the induction of hypertrophy in myotubes was conducted. To investigate whether AS promoted the hypertrophy of myotubes, an established in vitro model of myotube hypertrophy with and without AS was used and examined using microscopic images. The role of the PI3K/Akt/mTOR signaling pathway in AS-induced myotube hypertrophy was evaluated. Two inhibitors, wortmannin (an inhibitor of PI3K) and rapamycin (an inhibitor of mTOR), were used. Result The results revealed that the myotube diameters in the AS-treated group were significantly larger than those in the untreated control group (P < 0.05). Wortmannin and rapamycin inhibited AS-induced hypertrophy. Furthermore, AS increased Akt and mTOR phosphorylation through the PI3K pathway and induced myotube hypertrophy. Conclusion The results confirmed that AS induces hypertrophy in myotubes through the PI3K/Akt/mTOR pathway. PMID:24884709

  7. Phosphatidylinositol 3-kinase and calcium-activated transcription pathways are required for VLDL-induced smooth muscle cell proliferation.

    PubMed

    Lipskaia, Larissa; Pourci, Marie-Luce; Deloménie, Claudine; Combettes, Laurent; Goudounèche, Dominique; Paul, Jean-Louis; Capiod, Thierry; Lompré, Anne-Marie

    2003-05-30

    Little is known regarding the molecular mechanisms of atherogenicity of triglyceride-rich lipoproteins such as very low-density lipoproteins (VLDLs). We examined the effect of VLDL on proliferation of rat aortic smooth muscle cells, intracellular Ca2+ handling, and activity of cAMP-responsive element binding protein (CREB) and nuclear factor of activated T cells (NFAT) transcription factors. VLDL, isolated from human serum, dose- and time-dependently promoted proliferation. After 4 hours of exposure to VLDL (0.15 g/L proteins), the caffeine-induced Ca2+ release was inhibited and the IP3-sensitive Ca2+ release induced by ATP (10 micromol/L) was markedly prolonged. In quiescent cells, CREB was phosphorylated (pCREB) and NFAT was present in the cytosol, whereas in cells exposed to VLDL for 4 to 24 hours, pCREB disappeared and NFAT was translocated to the nucleus. VLDL-induced NFAT translocation and proliferation were blocked by cyclosporin A and LY294002 involving calcineurin and phosphatidylinositol 3-kinase (PI3K) pathways. Indeed, VLDLs rapidly phosphorylate protein kinase B and glycogen synthase kinase-3beta in a PI3K-dependent way. These results provide the first evidence that VLDLs induce smooth muscle cell proliferation by activating the PI3K pathway and nuclear NFAT translocation. Blockade of the Ca2+-induced Ca2+ release mechanism and dephosphorylation of pCREB contribute but were not sufficient to induce a proliferating phenotype.

  8. Static magnetic field enhances the viability and proliferation rate of adipose tissue-derived mesenchymal stem cells potentially through activation of the phosphoinositide 3-kinase/Akt (PI3K/Akt) pathway.

    PubMed

    Marędziak, Monika; Tomaszewski, Krzysztof; Polinceusz, Paulina; Lewandowski, Daniel; Marycz, Krzysztof

    2017-01-01

    The aim of this work was to investigate the effects of 0.5T static magnetic field (sMF) on the viability and proliferation rate of human adipose-derived mesenchymal stromal stem cells (hASCs) via activation of the phosphoinositide 3-kinase/Akt (PI3K/Akt) signaling pathway. In a 7-d culture we examined cell growth kinetic and population doubling time (PDT). We also examined cell morphology and the cellular senescence markers level. Exposure to sMF enhanced the viability of these cells. However, the effect was blocked by treating the cells with LY294002, a P13K inhibitor. We compared this effect by Western Blot analysis of Akt protein expression. We also examined whether the cell response on sMF stimulation is dependent on integrin engagement and we measured integrin gene expression. Our results suggest that stimulation using sMF is a viable method to improve hASC viability. sMF is involved in mechanisms associated with controlling cell proliferative potential signaling events.

  9. Inhibition of phosphatidylinositol 3-kinase promotes tumor cell resistance to chemotherapeutic agents via a mechanism involving delay in cell cycle progression

    SciTech Connect

    McDonald, Gail T.; Sullivan, Richard; Pare, Genevieve C.; Graham, Charles H.

    2010-11-15

    Approaches to overcome chemoresistance in cancer cells have involved targeting specific signaling pathways such as the phosphatidylinositol 3-kinase (PI3K) pathway, a stress response pathway known to be involved in the regulation of cell survival, apoptosis and growth. The present study determined the effect of PI3K inhibition on the clonogenic survival of human cancer cells following exposure to various chemotherapeutic agents. Treatment with the PI3K inhibitors LY294002 or Compound 15e resulted in increased survival of MDA-MB-231 breast carcinoma cells after exposure to doxorubicin, etoposide, 5-fluorouracil, and vincristine. Increased survival following PI3K inhibition was also observed in DU-145 prostate, HCT-116 colon and A-549 lung carcinoma cell lines exposed to doxorubicin. Increased cell survival mediated by LY294002 was correlated with a decrease in cell proliferation, which was linked to an increase in the proportion of cells in the G{sub 1} phase of the cell cycle. Inhibition of PI3K signaling also resulted in higher levels of the cyclin-dependent kinase inhibitors p21{sup Waf1/Cip1} and p27{sup Kip1}; and knockdown of p27{sup kip1} with siRNA attenuated resistance to doxorubicin in cells treated with LY294002. Incubation in the presence of LY294002 after exposure to doxorubicin resulted in decreased cell survival. These findings provide evidence that PI3K inhibition leads to chemoresistance in human cancer cells by causing a delay in cell cycle; however, the timing of PI3K inhibition (either before or after exposure to anti-cancer agents) may be a critical determinant of chemosensitivity.

  10. Apelin-13 promotes cardiomyocyte hypertrophy via PI3K-Akt-ERK1/2-p70S6K and PI3K-induced autophagy.

    PubMed

    Xie, Feng; Liu, Wei; Feng, Fen; Li, Xin; He, Lu; Lv, Deguan; Qin, Xuping; Li, Lifang; Li, Lanfang; Chen, Linxi

    2015-12-01

    Apelin is highly expressed in rat left ventricular hypertrophy Sprague Dawley rat models, and it plays a crucial role in the cardiovascular system. The aim this study was to clarify whether apelin-13 promotes hypertrophy in H9c2 rat cardiomyocytes and to investigate its underlying mechanism. The cardiomyocyte hypertrophy was observed by measuring the diameter, volume, and protein content of H9c2 cells. The activation of autophagy was evaluated by observing the morphology of autophagosomes by transmission electron microscopy, observing the subcellular localization of LC3 by light microscopy, and detecting the membrane-associated form of LC3 by western blot analysis. The phosphatidylinositol 3-kinase (PI3K) signaling pathway was identified and the proteins expression was detected using western blot analysis. The results revealed that apelin-13 increased the diameter, volume, and protein content of H9c2 cells and promoted the phosphorylation of PI3K, Akt, ERK1/2, and p70S6K. Apelin-13 activated the PI3K-Akt-ERK1/2-p70S6K pathway. PI3K inhibitor LY294002, Akt inhibitor 1701-1, ERK1/2 inhibitor PD98059 attenuated the increase of the cell diameter, volume, protein content induced by apelin-13. Apelin-13 increased the autophagosomes and up-regulated the expressions of beclin 1 and LC3-II/I both transiently and stably. The autophagy inhibitor 3MA ameliorated the increase of cell diameter, volume, and protein content that were induced by apelin-13. These results suggested that apelin-13 promotes H9c2 rat cardiomyocyte hypertrophy via PI3K-Akt-ERK1/2-p70S6K and PI3K-induced autophagy. © The Author 2015. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

  11. Strategically Timing Inhibition of Phosphatidylinositol 3-Kinase to Maximize Therapeutic Index in Estrogen Receptor Alpha-Positive, PIK3CA-Mutant Breast Cancer.

    PubMed

    Yang, Wei; Hosford, Sarah R; Dillon, Lloye M; Shee, Kevin; Liu, Stephanie C; Bean, Jennifer R; Salphati, Laurent; Pang, Jodie; Zhang, Xiaolin; Nannini, Michelle A; Demidenko, Eugene; Bates, Darcy; Lewis, Lionel D; Marotti, Jonathan D; Eastman, Alan R; Miller, Todd W

    2016-05-01

    Phosphatidylinositol 3-kinase (PI3K) inhibitors are being developed for the treatment of estrogen receptor α (ER)-positive breast cancer in combination with antiestrogens. Understanding the temporal response and pharmacodynamic effects of PI3K inhibition in ER(+) breast cancer will provide a rationale for treatment scheduling to maximize therapeutic index. Antiestrogen-sensitive and antiestrogen-resistant ER(+) human breast cancer cell lines and mice bearing PIK3CA-mutant xenografts were treated with the antiestrogen fulvestrant, the PI3K inhibitor GDC-0941 (pictilisib; varied doses/schedules that provided similar amounts of drug each week), or combinations. Cell viability, signaling pathway inhibition, proliferation, apoptosis, tumor volume, and GDC-0941 concentrations in plasma and tumors were temporally measured. Treatment with the combination of fulvestrant and GDC-0941, regardless of dose/schedule, was significantly more effective than that with single-agent treatments in fulvestrant-resistant tumors. Short-term, complete PI3K inhibition blocked cell growth in vitro more effectively than chronic, incomplete inhibition. Longer-term PI3K inhibition hypersensitized cells to growth factor signaling upon drug withdrawal. Different schedules of GDC-0941 elicited similar tumor responses. While weekly high-dose GDC-0941 with fulvestrant continuously suppressed PI3K signaling for 72 hours, inducing a bolus of apoptosis and inhibiting proliferation, PI3K reactivation upon GDC-0941 washout induced a proliferative burst. Fulvestrant with daily low-dose GDC-0941 metronomically suppressed PI3K for 6 to 9 hours/day, repeatedly inducing small amounts of apoptosis and temporarily inhibiting proliferation, followed by proliferative rebound compared with fulvestrant alone. Continuous and metronomic PI3K inhibition elicits robust anticancer effects in ER(+), PIK3CA-mutant breast cancer. Clinical exploration of alternate treatment schedules of PI3K inhibitors with antiestrogens

  12. [Silica induced α-SMA expression in HBE cell line by targeting the PI3K/Akt pathway].

    PubMed

    Li, Ai-ping; Hou, Zhi-guo; Fan, Jing-jing; Ji, Xiao-ming; Wang, Ting; Ni, Chun-hui

    2012-12-01

    To explore the role of the phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) pathway in silica-induced α-SMA (α smooth muscle actin) expression in HEB (human bronchial epithelial) cell. The cultured HBE cells were divided into 5 groups: control, silica, PI3K inhibitor (Ly294002), both PI3K inhibitor (Ly294002) and silica at the same time and the inhibitor 24 h ahead of silica. The final concentrations of PI3K inhibitor and silica were 10 µmol/L and 100 µg/ml, respectively. Western blots were used to detect protein expressions of Akt, p-Akt, TGF-β and α-SMA. The location and expression of α-SMA were measured by immunofluorescence assay. HBE cell line exposed to silica can induce Akt phosphorylation, in which expressions of p-Akt were up regulated 1 times at 48 and the highest at 72 h. The expressions of TGFβ increased remarkably at 12 h and the peak at 48 h after silica exposure, while the expressions of α-SMA increased at 24 h and the highest at 72 h. However, the PI3K inhibitor (Ly294002) significantly down regulated α-SMA expression. When the cell line exposed to the PI3K inhibitor ahead of silica 24 h, the expressions of p-Akt and α-SMA were more remarkably down regulated which were decreased 1.5 times and 7.6 times respectively compare to silica exposure group. But no significant changes were found for TGFβ expressions. The immunofluorescence assay showed that silica can induce α-SMA expression, which located in cytoplasma, and PI3K inhibitor can decrease the expression. Silica induced α-SMA expression in HBE cell line is by targeting the PI3K/Akt pathway and PI3K inhibitor can repress α-SMA expression.

  13. The PI3K Pathway: Background and Treatment Approaches.

    PubMed

    Lux, Michael P; Fasching, Peter A; Schrauder, Michael G; Hein, Alexander; Jud, Sebastian M; Rauh, Claudia; Beckmann, Matthias W

    2016-12-01

    Two-thirds of all breast cancer patients with metastases have a hormone receptor (HR)-positive/human epidermal growth factor receptor 2 (HER2)-negative subtype. Endocrine therapy is the treatment of choice in these patients since in addition to its effectiveness it can also maintain the patients' quality of life over a longer term. However, 44-62% of postmenopausal patients with metastatic breast carcinoma have primary tamoxifen resistance. After 3-5 years, 30-40% of the patients receiving tamoxifen treatment develop secondary resistance. Understanding the way in which resistance develops is therefore essential for developing treatment approaches that can prevent or reverse endocrine resistance. The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway plays a central role here. As a result of the numerous interactions involved, complex issues arise that need to be taken into account in the development and use of therapeutic agents. In addition, this signaling pathway is the one that most frequently undergoes mutations in breast cancer. The prognostic and predictive significance of individual mutations has not yet been fully explained, but it might provide a basis for patient selection in clinical studies. Initial research results on the use of PI3K inhibitors suggest that this may be a highly promising therapeutic approach, with an acceptable side effect profile.

  14. The phosphatidylinositol-3 kinase/Akt pathway mediates geranylgeranylacetone-induced neuroprotection against cerebral infarction in rats.

    PubMed

    Abe, Eiji; Fujiki, Minoru; Nagai, Yasuyuki; Shiqi, Kong; Kubo, Takeshi; Ishii, Keisuke; Abe, Tatsuya; Kobayashi, Hidenori

    2010-05-12

    Previous studies demonstrated the cytoprotective effect of geranylgeranylacetone (GGA), a heat shock protein inducer, against ischemic insult. Phosphatidylinositol-3 kinase/Akt (PI3K/Akt) is thought to be an important factor that mediates neuroprotection. However, the signaling pathways in the brain in vivo after oral GGA administration remain unclear. We measured and compared infarction volumes to investigate the effect of GGA on cerebral infarction induced by permanent middle cerebral artery occlusion in rats. We evaluated the effects of pretreatment with 5-hydroxydecanoate (5HD), a specific mitochondrial ATP-sensitive potassium (mitoK(ATP)) channel inhibitor; diazoxide (DZX), a selective mitoK(ATP) channel opener and wortmannin (Wort), a specific PI3K inhibitor of GGA-induced neuroprotection against infarction volumes. To clarify the relationship between PI3K/Akt activation and neuroprotection, we used immunoblot analysis to determine the amount of p-Akt proteins present after GGA administration with or without Wort treatment. Neuroprotective effects of GGA (pretreatment with a single oral GGA dose (800 mg/kg) 48 h before ischemia) were prevented by 5HD, DZX and Wort pretreatment, which indicates that the selective mitoK(ATP) channel and the PI3K/Akt pathway may mediate GGA-dependent protection. Oral GGA-induced p-Akt and GGA pretreatment enhanced ischemia-induced p-Akt, both of which were prevented by Wort pretreatment. These results suggest that a single oral dose of GGA induces p-Akt and that GGA plays an important role in neuroprotection against cerebral ischemia through the mitoK(ATP) channel opening. (c) 2010 Elsevier B.V. All rights reserved.

  15. Low [Mg2+]e enhances arterial spontaneous tone via phosphatidylinositol 3-kinase in DOCA-salt hypertension.

    PubMed

    Northcott, Carrie A; Watts, Stephanie W

    2004-01-01

    Phosphatidylinositol 3-kinase (PI3K) has been implicated in low extracellular Mg2+ concentration ( [Mg2+]e)-induced aortic contraction, and Mg2+ deficiency has been associated with hypertension. Moreover, arterial PI3K activity is increased in hypertensive deoxycorticosterone (DOCA)-salt rats. We hypothesized that low [Mg2+]e activates PI3K, eliciting enhanced vascular contraction, PI3K activity, and norepinephrine (NE)-induced contraction. Spontaneous tone was monitored in endothelium-denuded aortic strips from sham and DOCA-salt rats exposed to low Mg2+ (0.15 mmol/L), high Mg2+ (4.8 mmol/L), or normal (1.17 mmol/L) physiologic salt solution (PSS) in isolated tissue baths. LY294002 (20 micromol/L), a PI3K inhibitor, or vehicle was added (30 minutes), followed by NE (10(-9) to 3 x10(-5) mol/L). Low [Mg2+]e significantly enhanced tone in aortas from DOCA-salt and sham rats compared with normal PSS (DOCA-salt low [Mg2+]e, +51.5 +7.0 vs DOCA-salt normal PSS, +7.1 +1.4 % of initial phenylephrine [PE] contraction). LY294002 and incubation with high Mg2+ PSS decreased tone in aortas from DOCA-salt rats (low [Mg2+]e LY294002, --87.5 +8.8; normal PSS LY294002, -81.7 +13.7; and high [Mg2+]e, -31.2 +10.8 % of initial PE contraction). Low [Mg2+]e leftward-shifted NE-induced aortic contractions in sham and thus matched the shift observed with DOCA (-log EC50 mol/L: sham PSS, -7.7 +0.1; DOCA-salt PSS, -8.2 +0.1; sham low [Mg2+]e, -8.2 +0.1; and DOCA-salt low [Mg2+]e, -8.1 +0.1). Moreover, this shift was inhibited by LY294002. In conclusion, low [Mg2+]e might activate PI3K, leading to enhanced tone and agonist-induced contraction observed in aortas from DOCA-salt hypertensive rats.

  16. Carbamazepine enhances the activity of glutamate transporter type 3 via phosphatidylinositol 3-kinase.

    PubMed

    Lee, Gwanwoo; Huang, Yueming; Washington, Jacqueline M; Briggs, Nicole W; Zuo, Zhiyi

    2005-01-01

    Glutamate transporters (also called excitatory amino acid transporters, EAAT) participate in maintaining extracellular homeostasis of glutamate, a major excitatory neurotransmitter, and regulating glutamate neurotransmission. EAAT3, the major neuronal EAAT, may also regulate gamma-aminobutyric acid-mediated inhibitory neurotransmission. Dysfunction of EAAT3 has been shown to induce seizure in rats. We hypothesize that carbamazepine, a commonly used antiepileptic agent, enhances EAAT3 activity. We tested this hypothesis using oocytes artificially expressing EAAT3 and C6 rat glioma cells expressing endogenous EAAT3. In oocytes, carbamazepine dose-dependently enhanced EAAT3 activity. The EC50 of this carbamazepine effect was 12.2muM. The concentrations of carbamazepine to significantly enhance EAAT3 activity were within the therapeutic serum levels (17-51muM) of carbamazepine for the antiepileptic effect. Carbamazepine decreased the Km but did not change the maximal response of EAAT3 to glutamate. Carbamazepine-increased EAAT3 activity was inhibited by wortmannin or LY-294002, phosphatidylinositol 3-kinase (PI3K) inhibitors, but was not affected by staurosporine, chelerythrine or calphostin C, protein kinase C inhibitors. In C6 cells, carbamazepine also enhanced the endogenous EAAT3 activity. However, carbamazepine did not affect the activity of EAAT4 expressed in Cos7 cells. These results suggest that carbamazepine at clinically relevant concentrations specifically enhances the affinity of EAAT3 for glutamate to increase EAAT3 activity via a PI3K-dependent pathway. EAAT3 may be a therapeutic target for carbamazepine in the central nervous system.

  17. Oxidative stress stimulates skeletal muscle glucose uptake through a phosphatidylinositol 3-kinase-dependent pathway

    PubMed Central

    Higaki, Yasuki; Mikami, Toshio; Fujii, Nobuharu; Hirshman, Michael F.; Koyama, Katsuhiro; Seino, Tetsuya; Tanaka, Keitaro; Goodyear, Laurie J.

    2010-01-01

    We determined the acute effects of oxidative stress on glucose uptake and intracellular signaling in skeletal muscle by incubating muscles with reactive oxygen species (ROS). Xanthine oxidase (XO) is a superoxide-generating enzyme that increases ROS. Exposure of isolated rat extensor digitorum longus (EDL) muscles to Hx/XO (Hx/XO) for 20 min resulted in a dose-dependent increase in glucose uptake. To determine whether the mechanism leading to Hx/XO-stimulated glucose uptake is associated with the production of H2O2, EDL muscles from rats were preincubated with the H2O2 scavenger catalase or the superoxide scavenger superoxide dismutase (SOD) prior to incubation with Hx/XO. Catalase treatment, but not SOD, completely inhibited the increase in Hx/XO-stimulated 2-deoxyglucose (2-DG) uptake, suggesting that H2O2 is an intermediary leading to Hx/XO-stimulated glucose uptake with incubation. Direct H2O2 also resulted in a dose-dependent increase in 2-DG uptake in isolated EDL muscles, and the maximal increase was threefold over basal levels at a concentration of 600 μmol/l H2O2. H2O2-stimulated 2-DG uptake was completely inhibited by the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin, but not the nitric oxide inhibitor NG-monomethyl-L-arginine. H2O2 stimulated the phosphorylation of Akt Ser473 (7-fold) and Thr308 (2-fold) in isolated EDL muscles. H2O2 at 600 μmol/l had no effect on ATP concentrations and did not increase the activities of either the α1 or α2 catalytic isoforms of AMP-activated protein kinase. These results demonstrate that acute exposure of muscle to ROS is a potent stimulator of skeletal muscle glucose uptake and that this occurs through a PI3K-dependent mechanism. PMID:18303121

  18. Down-regulation of PKHD1 induces cell apoptosis through PI3K and NF-{kappa}B pathways

    SciTech Connect

    Sun, Liping; Wang, Shixuan; Hu, Chaofeng; Zhang, Xinzhou

    2011-04-15

    Mutations in PKHD1 (polycystic kidney and hepatic disease gene 1) gene cause the autosomal recessive polycystic kidney disease (ARPKD). Fibrocystin/polyductin (FPC), encoded by PKHD1, is a membrane-associated receptor-like protein. Although it is widely accepted that cystogenesis is mostly due to aberrant cell proliferation and apoptosis, it is still unclear how apoptosis is regulated. The aim of this study is to analyze the relationship among apoptosis, phosphatidylinositol 3-kinase (PI3K)/Akt and nuclear factor {kappa}B (NF-{kappa}B) in FPC knockdown kidney cells. We show that PKHD1-silenced HEK293 cells demonstrate a higher PI3K/Akt activity. Selective inhibition of PI3K/Akt using LY294002 or wortmannin in these cells increases serum starvation-induced HEK293 cell apoptosis with a concomitant decrease in cell proliferation and higher caspase-3 activity. PI3K/Akt inhibition also leads to increased NF-{kappa}B activity in these cells. We conclude that the PI3K/Akt pathway is involved in apoptotic function in PKHD1-silenced cells, and PI3K/Akt inhibition correlates with upregulation of NF-{kappa}B activity. These observations provide a potential platform for determining FPC function and therapeutic investigation of ARPKD.

  19. Phosphatidylinositol-3-kinase as a putative target for anticancer action of clotrimazole.

    PubMed

    Furtado, Cristiane M; Marcondes, Mariah C; Carvalho, Renato S; Sola-Penna, Mauro; Zancan, Patricia

    2015-05-01

    Clotrimazole (CTZ) has been proposed as an antitumoral agent because of its properties that inhibit glycolytic enzymes and detach them from the cytoskeleton. However, the broad effects of the drug, e.g., acting on different enzymes and pathways, indicate that CTZ might also affect several signaling pathways. In this study, we show that CTZ interferes with the human breast cancer cell line MCF-7 after a short incubation period (4 h), thereby diminishing cell viability, promoting apoptosis, depolarizing mitochondria, inhibiting key glycolytic regulatory enzymes, decreasing the intracellular ATP content, and permeating plasma membranes. CTZ treatment also interferes with autophagy. Moreover, when the incubation is performed under hypoxic conditions, certain effects of CTZ are enhanced, such as phosphatidylinositol-3-phosphate kinase (PI3K), which is inhibited upon CTZ treatment; this inhibition is potentiated under hypoxia. CTZ-induced PI3K inhibition is not caused by upstream effects of CTZ because the drug does not affect the interaction of the PI3K regulatory subunit and the insulin receptor substrate (IRS)-1. Additionally, CTZ directly inhibits human purified PI3K in a dose-dependent and reversible manner. Pharmacologic and in silico results suggest that CTZ may bind to the PI3K catalytic site. Therefore, we conclude that PI3K is a novel, putative target for the antitumoral effects of CTZ, interfering with autophagy, apoptosis, cell division and viability.

  20. Molecular cloning and biochemical characterization of a Drosophila phosphatidylinositol-specific phosphoinositide 3-kinase.

    PubMed

    Linassier, C; MacDougall, L K; Domin, J; Waterfield, M D

    1997-02-01

    Molecular, biochemical and genetic characterization of phosphoinositide 3-kinases (PI3Ks) have identified distinct classes of enzymes involved in processes mediated by activation of cell-surface receptors and in constitutive intracellular protein trafficking events. The latter process appears to involve a PtdIns-specific PI3K first described in yeast as a mutant, vps34, defective in the sorting of newly synthesized proteins from the Golgi to the vacuole. We have identified a representative member of each class of PI3Ks in Drosophila using a PCR-based approach. In the present paper we describe the molecular cloning of a PI3K from Drosophila, P13K_59F, that shows sequence similarity to Vps34. PI3K_59F encodes a protein of 108 kDa co-linear with Vps34 homologues, and with three regions of sequence similarity to other PI3Ks. Biochemical characterization of the enzyme, by expression of the complete coding sequence as a glutathione S-transferase fusion protein in Sf9 cells, demonstrates that PI3K_59F is a PtdIns-specific PI3K that can utilize either Mg2+ or Mn2+. This activity is sensitive to inhibition both by non-ionic detergent (Nonidet P40) and by wortmannin (IC50 10 nM). PI3K_59F, therefore, conserves both the structural and biochemical properties of the Vps34 class of enzymes.

  1. Autophagy inhibition enhances colorectal cancer apoptosis induced by dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor NVP-BEZ235

    PubMed Central

    YANG, XIAOYU; NIU, BINGXUAN; WANG, LIBO; CHEN, MEILING; KANG, XIAOCHUN; WANG, LUONAN; JI, YINGHUA; ZHONG, JIATENG

    2016-01-01

    Phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) signaling pathway performs a central role in tumorigenesis and is constitutively activated in many malignancies. As a novel dual PI3K/mTOR inhibitor currently undergoing evaluation in a phase I/II clinical trial, NVP-BEZ235 indicates a significant antitumor efficacy in diverse solid tumors, including colorectal cancer (CRC). Autophagy is a catabolic process that maintains cellular homeostasis and reduces diverse stresses through lysosomal recycling of the unnecessary and damaged cell components. This process is also observed to antagonize the antitumor efficacy of PI3K/mTOR inhibitor agents such as NVP-BEZ235, via apoptosis inhibition. In the present study, we investigated anti-proliferative and apoptosis-inducing ability of NVP-BEZ235 in SW480 cells and the crosstalk between autophagy and apoptosis in SW480 cells treated with NVP-BEZ235 in combination with an autophagy inhibitor. The results revealed that, NVP-BEZ235 effectively inhibit the growth of SW480 cells by targeting the PI3K/mTOR signaling pathway and induced apoptosis. The inhibition of autophagy with 3-methyladenine or chloroquine inhibitors in combination with NVP-BEZ235 in SW480 cells enhanced the apoptotic rate as componets to NVP-BEZ235 alone. In conclusion, the findings provide a rationale for chemotherapy targeting the PI3K/mTOR signaling pathway presenting a potential therapeutic strategy to enhance the efficacy of dual PI3K/mTOR inhibitor NVP-BEZ235 in combination with an autophagy inhibitor in CRC treatment and treatment of other tumors. PMID:27347108

  2. High-Throughput Small Interfering RNA Screening Identifies Phosphatidylinositol 3-Kinase Class II Alpha as Important for Production of Human Cytomegalovirus Virions

    PubMed Central

    Polachek, William S.; Moshrif, Hanan F.; Franti, Michael; Coen, Donald M.; Sreenu, Vattipally B.

    2016-01-01

    ABSTRACT High-throughput small interfering RNA (siRNA) screening is a useful methodology to identify cellular factors required for virus replication. Here we utilized a high-throughput siRNA screen based on detection of a viral antigen by microscopy to interrogate cellular protein kinases and phosphatases for their importance during human cytomegalovirus (HCMV) replication and identified the class II phosphatidylinositol 3-kinase class II alpha (PI3K-C2A) as being involved in HCMV replication. Confirming this observation, infected cells treated with either pooled or individual siRNAs targeting PI3K-C2A mRNA produced approximately 10-fold less infectious virus than the controls. Western blotting and quantitative PCR analysis of infected cells treated with siRNAs indicated that depletion of PI3K-C2A slightly reduced the accumulation of late but not immediate early or early viral antigens and had no appreciable effect on viral DNA synthesis. Analysis of siRNA-treated cells by electron microscopy and Western blotting indicated that PI3K-C2A was not required for the production of viral capsids but did lead to increased numbers of enveloped capsids in the cytoplasm that had undergone secondary envelopment and a reduction in the amount of viral particles exiting the cell. Therefore, PI3K-C2A is a factor important for HCMV replication and has a role in the production of HCMV virions. IMPORTANCE There is limited information about the cellular factors required for human cytomegalovirus (HCMV) replication. Therefore, to identify proteins involved in HCMV replication, we developed a methodology to conduct a high-throughput siRNA screen of HCMV-infected cells. From our screening data, we focused our studies on the top hit from our screen, the lipid kinase phosphatidylinositol 3-kinase class II alpha (PI3K-C2A), as its role in HCMV replication was unknown. Interestingly, we found that PI3K-C2A is important for the production of HCMV virions and is involved in virion production

  3. Inhibition of the PI3K Pathway Suppresses Hormonal Secretion and Limits Growth in Pheochromocytoma Cells

    PubMed Central

    Adler, Joel T.; Hottinger, Daniel G.

    2009-01-01

    Background Operative resection is the only curative treatment for pheochromocytomas. Inhibition of the phosphatidylinositol-3 kinase (PI3K)-Akt pathway has been shown to be an effective treatment of neuroendocrine (NE) tumors in vitro. We hypothesized that inhibition of the PI3K-Akt pathway would be a viable strategy to inhibit growth and hormonal secretion in pheochromocytoma cells. Methods Sixteen pheochromocytomas were analyzed for expression of phosphorylated Akt and the NE marker achaete scute complex-like 1 (ASCL1). Pheochromocytoma PC-12 cells were treated with up to 100 µM of the PI3K-specific inhibitor LY294002 for 48 h. Western blot analysis was used to measure phosphorylated Akt, total Akt, ASCL1, chromogranin A (CgA), and markers of apoptosis. Growth was assessed by a methylthiazolyldiphenyl-tetrazolium (MTT) bromide cellular proliferation assay for six days. Results Human pheochromocytomas expressed significant amounts of phosphorylated Akt, and there was a significant correlation between malignant pheochromocytomas and the amount of expressed ASCL1. LY294002 significantly inhibited the PI3K-Akt pathway. Treatment led to a dose-dependent decrease in both ASCL1 and CgA, indicating an alteration in the NE phenotype and hormonal suppression. Treatment decreased cellular proliferation, and cleavage of the apoptotic markers caspase-3 and PARP was observed. Conclusions Human pheochromocytoma tumor samples express high levels of phosphorylated Akt. LY294002 effectively inhibits the PI3K-Akt pathway, suppresses NE tumor markers, and decreases cellular proliferation via apoptosis in vitro. Inhibition of the PI3K pathway may represent a new strategy in the treatment of pheochromocytomas. PMID:19669229

  4. The Emerging Role of the Phosphatidylinositol 3-Kinase/ Akt/Mammalian Target of Rapamycin Signaling Network in Cancer Stem Cell Biology

    PubMed Central

    Martelli, Alberto M.; Evangelisti, Camilla; Chiarini, Francesca; Grimaldi, Cecilia; McCubrey, James A.

    2010-01-01

    The cancer stem cell theory entails the existence of a hierarchically organized, rare population of cells which are responsible for tumor initiation, self-renewal/maintenance, and mutation accumulation. The cancer stem cell proposition could explain the high frequency of cancer relapse and resistance to currently available therapies. The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway regulates a wide array of physiological cell functions which include differentiation, proliferation, survival, metabolism, autophagy, and motility. Dysregulated PI3K/Akt/mTOR signaling has been documented in many types of neoplasias. It is now emerging that this signaling network plays a key role in cancer stem cell biology. Interestingly, cancer stem cells displayed preferential sensitivity to pathway inhibition when compared to healthy stem cells. This observation provides the proof-of-principle that functional differences in signaling pathways between neoplastic stem cells and healthy stem cells could be identified. In this review, we present the evidence which links the signals emanating from the PI3K/Akt/mTOR cascade with the functions of cancer stem cells, both in solid and hematological tumors. We then highlight how targeting PI3K/Akt/mTOR signaling with small molecules could improve cancer patient outcome. PMID:24281174

  5. miR-362-5p inhibits proliferation and migration of neuroblastoma cells by targeting phosphatidylinositol 3-kinase-C2β.

    PubMed

    Wu, Kai; Yang, Liucheng; Chen, Jianfeng; Zhao, Haijun; Wang, Jianjun; Xu, Shuai; Huang, Zonghai

    2015-07-08

    miR-362-5p is down-regulated in high-risk neuroblastoma and can function as a tumor suppressor. However, its role remains poorly understood. We show that miR-362-5p is down-regulated in metastatic neuroblastoma compared with primary neuroblastoma. Overexpression of miR-362-5p inhibits cell proliferation, migration and invasion of neuroblastoma cells in vitro and suppresses tumor growth of neuroblastoma in vivo. Phosphatidylinositol 3-kinase (PI3K)-C2β is a target of miR-362-5p. Knockdown of PI3K-C2β by siRNA had a similar effect to overexpression of miR-362-5p on SH-SY5Y cells. Overexpression of PI3K-C2β partially reversed tumor-suppressive effects of miR-362-5p. We suggest that miR-362-5p suppresses neuroblastoma cell growth and motility, partially by targeting PI3K-C2β.

  6. The Role of the PI3K Signaling Pathway in CD4+ T Cell Differentiation and Function

    PubMed Central

    Han, Jonathan M.; Patterson, Scott J.; Levings, Megan K.

    2012-01-01

    The relative activity of regulatory versus conventional CD4+ T cells ultimately maintains the delicate balance between immune tolerance and inflammation. At the molecular level, the activity of phosphatidylinositol 3-kinase (PI3K) and its downstream positive and negative regulators has a major role in controlling the balance between immune regulation and activation of different subsets of effector CD4+ T cells. In contrast to effector T cells which require activation of the PI3K to differentiate and mediate their effector function, regulatory T cells rely on minimal activation of this pathway to develop and maintain their characteristic phenotype, function, and metabolic state. In this review, we discuss the role of the PI3K signaling pathway in CD4+ T cell differentiation and function, and focus on how modulation of this pathway in T cells can alter the outcome of an immune response, ultimately tipping the balance between tolerance and inflammation. PMID:22905034

  7. PDK1 controls upstream PI3K expression and PIP3 generation.

    PubMed

    Dieterle, A M; Böhler, P; Keppeler, H; Alers, S; Berleth, N; Drießen, S; Hieke, N; Pietkiewicz, S; Löffler, A S; Peter, C; Gray, A; Leslie, N R; Shinohara, H; Kurosaki, T; Engelke, M; Wienands, J; Bonin, M; Wesselborg, S; Stork, B

    2014-06-05

    The PI3K/PDK1/Akt signaling axis is centrally involved in cellular homeostasis and controls cell growth and proliferation. Due to its key function as regulator of cell survival and metabolism, the dysregulation of this pathway is manifested in several human pathologies including cancers and immunological diseases. Thus, current therapeutic strategies target the components of this signaling cascade. In recent years, numerous feedback loops have been identified that attenuate PI3K/PDK1/Akt-dependent signaling. Here, we report the identification of an additional level of feedback regulation that depends on the negative transcriptional control of phosphatidylinositol 3-kinase (PI3K) class IA subunits. Genetic deletion of 3-phosphoinositide-dependent protein kinase 1 (PDK1) or the pharmacological inhibition of its downstream effectors, that is, Akt and mammalian target of rapamycin (mTOR), relieves this suppression and leads to the upregulation of PI3K subunits, resulting in enhanced generation of phosphatidylinositol-3,4,5-trisphosphate (PIP3). Apparently, this transcriptional induction is mediated by the concerted action of different transcription factor families, including the transcription factors cAMP-responsive element-binding protein and forkhead box O. Collectively, we propose that PDK1 functions as a cellular sensor that balances basal PIP3 generation at levels sufficient for survival but below a threshold being harmful to the cell. Our study suggests that the efficiency of therapies targeting the aberrantly activated PI3K/PDK1/Akt pathway might be increased by the parallel blockade of feedback circuits.

  8. Deoxycholyltaurine Rescues Human Colon Cancer Cells From Apoptosis by Activating EGFR-Dependent PI3K/Akt Signaling

    PubMed Central

    Raufman, Jean-Pierre; Shant, Jasleen; Guo, Chang Yue; Roy, Sanjit; Cheng, Kunrong

    2010-01-01

    Recent studies indicate that secondary bile acids promote colon cancer cell proliferation but their role in maintaining cell survival has not been explored. We found that deoxycholyltaurine (DCT) markedly attenuated both unstimulated and TNF-α-stimulated programmed cell death in colon cancer cells by a phosphatidylinositol 3-kinase (PI3K)-dependent mechanism. To examine the role of bile acids and PI3K signaling in maintaining colon cancer cell survival, we explored the role of signaling downstream of bile acid-induced activation of the epidermal growth factor receptor (EGFR) in regulating both apoptosis and proliferation of HT-29 and H508 human colon cancer cells. DCT caused dose- and time-dependent Akt (Ser473) phosphorylation, a commonly used marker of activated PI3K/Akt signaling. Both EGFR kinase and PI3K inhibitors attenuated DCT-induced Akt phosphorylation and Akt activation, as demonstrated by reduced phosphorylation of a GSK-3-paramyosin substrate. Transfection of HT-29 cells with kinase-dead EGFR (K721M) reduced DCT-induced Akt phosphorylation. In HT-29 cells, EGFR and PI3K inhibitors as well as transfection with dominant negative AKT attenuated DCT-induced cell proliferation. DCT-induced PI3K/Akt activation resulted in downstream phosphorylation of GSK-3 (Ser21/9) and BAD (Ser136), and nuclear translocation (activation) of NF-κB, thereby confirming that DCT-induced activation of PI3K/Akt signaling regulates both proproliferative and prosurvival signals. Collectively, these results indicate that DCT-induced activation of post-EGFR PI3K/Akt signaling stimulates both colon cancer cell survival and proliferation. PMID:18064605

  9. Effect of saw palmetto extract on PI3K cell signaling transduction in human glioma.

    PubMed

    Yang, Yang; Hui, Lv; Yuqin, Che; Jie, Li; Shuai, Hou; Tiezhu, Zhou; Wei, Wang

    2014-08-01

    Saw palmetto extract can induce the apoptosis of prostate cancer cells. The aim of the present study was to investigate the effect of saw palmetto extract on the phosphatidylinositol 3-kinase (PI3K)/Akt signaling transduction pathway in human glioma U87 and U251 cell lines. Suspensions of U87 and U251 cells in a logarithmic growth phase were seeded into six-well plates at a density of 10(4) cells/well. In the experimental group, 1 μl/ml saw palmetto extract was added, while the control group was cultured without a drug for 24 h. The expression levels of PI3K, B-cell lymphoma-extra large (Bcl-xL) and p53 were evaluated through western blot analysis. In the experimental group, the U87 and U251 cells exhibited a lower expression level of PI3K protein as compared with the control group (t=6.849; P<0.001). In addition, the two cell lines had a higher expression level of p53 protein in the experimental group as compared with the control group (t=40.810; P<0.001). Protein expression levels of Bcl-xL decreased significantly in the experimental group as compared with the control group (t=19.640; P=0.000). Therefore, saw palmetto extract induces glioma cell growth arrest and apoptosis via decreasing PI3K/Akt signal transduction.

  10. Regulation of the PI3K pathway through a p85α monomer–homodimer equilibrium

    PubMed Central

    Cheung, Lydia WT; Walkiewicz, Katarzyna W; Besong, Tabot MD; Guo, Huifang; Hawke, David H; Arold, Stefan T; Mills, Gordon B

    2015-01-01

    The canonical action of the p85α regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is to associate with the p110α catalytic subunit to allow stimuli-dependent activation of the PI3K pathway. We elucidate a p110α-independent role of homodimerized p85α in the positive regulation of PTEN stability and activity. p110α-free p85α homodimerizes via two intermolecular interactions (SH3:proline-rich region and BH:BH) to selectively bind unphosphorylated activated PTEN. As a consequence, homodimeric but not monomeric p85α suppresses the PI3K pathway by protecting PTEN from E3 ligase WWP2-mediated proteasomal degradation. Further, the p85α homodimer enhances the lipid phosphatase activity and membrane association of PTEN. Strikingly, we identified cancer patient-derived oncogenic p85α mutations that target the homodimerization or PTEN interaction surface. Collectively, our data suggest the equilibrium of p85α monomer–dimers regulates the PI3K pathway and disrupting this equilibrium could lead to disease development. DOI: http://dx.doi.org/10.7554/eLife.06866.001 PMID:26222500

  11. PI3K regulates BMAL1/CLOCK-mediated circadian transcription from the Dbp promoter.

    PubMed

    Morishita, Yoshikazu; Miura, Daiki; Kida, Satoshi

    2016-06-01

    The circadian rhythm generated by circadian clock underlies a molecular mechanism of rhythmic transcriptional regulation by transcription factor BMAL1/CLOCK. Importantly, the circadian clock is coordinated by exogenous cues to accommodate to changes in the external environment. However, the molecular mechanisms by which intracellular-signaling pathways mediate the adjustments of the circadian transcriptional rhythms remain unclear. In this study, we found that pharmacological inhibition or shRNA-mediated knockdown of phosphatidylinositol 3-kinase (PI3K) blocked upregulation of Dbp mRNA induced by serum shock in NIH 3T3 cells. Moreover, the inhibition of PI3K significantly reduced the promoter activity of the Dbp gene, as well as decreased the recruitment of BMAL1/CLOCK to the E-box in the Dbp promoter. Interestingly, the inhibition of PI3K blocked heterodimerization of BMAL1 and CLOCK. Our findings suggest that PI3K signaling plays a modulatory role in the regulation of the transcriptional rhythm of the Dbp gene by targeting BMAL1 and CLOCK.

  12. The PI3K/Akt pathway contributes to arenavirus budding.

    PubMed

    Urata, Shuzo; Ngo, Nhi; de la Torre, Juan Carlos

    2012-04-01

    Several arenaviruses, chiefly Lassa virus (LASV), cause hemorrhagic fever (HF) disease in humans and pose a significant public health concern in regions where they are endemic. On the other hand, evidence indicates that the globally distributed prototypic arenavirus lymphocytic choriomeningitis virus (LCMV) is a neglected human pathogen. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway participates in many cellular processes, including cell survival and differentiation, and also has been shown to play important roles in different steps of the life cycles of a variety of viruses. Here we report that the inhibition of the PI3K/Akt pathway inhibited budding and to a lesser extent RNA synthesis, but not cell entry, of LCMV. Accordingly, BEZ-235, a PI3K inhibitor currently in cancer clinical trials, inhibited LCMV multiplication in cultured cells. These findings, together with those previously reported for Junin virus (JUNV), indicate that targeting the PI3K/Akt pathway could represent a novel antiviral strategy to combat human-pathogenic arenaviruses.

  13. Effect of saw palmetto extract on PI3K cell signaling transduction in human glioma

    PubMed Central

    YANG, YANG; HUI, LV; YUQIN, CHE; JIE, LI; SHUAI, HOU; TIEZHU, ZHOU; WEI, WANG

    2014-01-01

    Saw palmetto extract can induce the apoptosis of prostate cancer cells. The aim of the present study was to investigate the effect of saw palmetto extract on the phosphatidylinositol 3-kinase (PI3K)/Akt signaling transduction pathway in human glioma U87 and U251 cell lines. Suspensions of U87 and U251 cells in a logarithmic growth phase were seeded into six-well plates at a density of 104 cells/well. In the experimental group, 1 μl/ml saw palmetto extract was added, while the control group was cultured without a drug for 24 h. The expression levels of PI3K, B-cell lymphoma-extra large (Bcl-xL) and p53 were evaluated through western blot analysis. In the experimental group, the U87 and U251 cells exhibited a lower expression level of PI3K protein as compared with the control group (t=6.849; P<0.001). In addition, the two cell lines had a higher expression level of p53 protein in the experimental group as compared with the control group (t=40.810; P<0.001). Protein expression levels of Bcl-xL decreased significantly in the experimental group as compared with the control group (t=19.640; P=0.000). Therefore, saw palmetto extract induces glioma cell growth arrest and apoptosis via decreasing PI3K/Akt signal transduction. PMID:25009620

  14. The PTEN/PI3K/AKT Pathway in vivo, Cancer Mouse Models

    PubMed Central

    Carnero, Amancio; Paramio, Jesus M.

    2014-01-01

    When PI3K (phosphatidylinositol-3 kinase) is activated by receptor tyrosine kinases, it phosphorylates PIP2 to generate PIP3 and activates the signaling pathway. Phosphatase and tensin homolog deleted on chromosome 10 dephosphorylates PIP3 to PIP2, and thus, negatively regulates the pathway. AKT (v-akt murine thymoma viral oncogene homolog; protein kinase B) is activated downstream of PIP3 and mediates physiological processes. Furthermore, substantial crosstalk exists with other signaling networks at all levels of the PI3K pathway. Because of its diverse array, gene mutations, and amplifications and also as a consequence of its central role in several signal transduction pathways, the PI3K-dependent axis is frequently activated in many tumors and is an attractive therapeutic target. The preclinical testing and analysis of these novel therapies requires appropriate and well-tailored systems. Mouse models in which this pathway has been genetically modified have been essential in understanding the role that this pathway plays in the tumorigenesis process. Here, we review cancer mouse models in which the PI3K/AKT pathway has been genetically modified. PMID:25295225

  15. Neuroprotection of brain-derived neurotrophic factor against hypoxic injury in vitro requires activation of extracellular signal-regulated kinase and phosphatidylinositol 3-kinase.

    PubMed

    Sun, Xiaomei; Zhou, Hui; Luo, Xiaoli; Li, Shengfu; Yu, Dan; Hua, Jiping; Mu, Dezhi; Mao, Meng

    2008-01-01

    Intrauterine asphyxia is one of the major contributors for perinatal death, mental and physical disorders of surviving children. Brain-derived neurotrophic factor (BDNF) provides a promising solution to hypoxic injury due to its survival-promoting effects. In an attempt to identify possible molecular mechanisms underlying the neuroprotective role of BDNF, we studied extracellular signal-regulated kinase (ERK), phosphatidylinositol 3-kinase (PI-3-K) and p38 mitogen-activated protein kinase (MAPK) pathways. We demonstrated that BDNF protected cortical neurons against hypoxic injury in vitro via activation of both the ERK and PI-3-K pathways but not the p38 MAPK pathway. We also showed that both hypoxic stimuli and exogenous BDNF treatment phosphorylated the cyclic AMP response element-binding protein (CREB) and that CREB phosphorylation induced by BDNF was mediated via the ERK pathway in cultured cortical neurons.

  16. The PI3K/Akt Pathway in Tumors of Endocrine Tissues

    PubMed Central

    Robbins, Helen Louise; Hague, Angela

    2016-01-01

    The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is a key driver in carcinogenesis. Defects in this pathway in human cancer syndromes such as Cowden’s disease and Multiple Endocrine Neoplasia result in tumors of endocrine tissues, highlighting its importance in these cancer types. This review explores the growing evidence from multiple animal and in vitro models and from analysis of human tumors for the involvement of this pathway in the following: thyroid carcinoma subtypes, parathyroid carcinoma, pituitary tumors, adrenocortical carcinoma, phaeochromocytoma, neuroblastoma, and gastroenteropancreatic neuroendocrine tumors. While data are not always consistent, immunohistochemistry performed on human tumor tissue has been used alongside other techniques to demonstrate Akt overactivation. We review active Akt as a potential prognostic marker and the PI3K pathway as a therapeutic target in endocrine neoplasia. PMID:26793165

  17. PI3K{gamma} activation by CXCL12 regulates tumor cell adhesion and invasion

    SciTech Connect

    Monterrubio, Maria; Mellado, Mario; Carrera, Ana C.

    2009-10-16

    Tumor dissemination is a complex process, in which certain steps resemble those in leukocyte homing. Specific chemokine/chemokine receptor pairs have important roles in both processes. CXCL12/CXCR4 is the most commonly expressed chemokine/chemokine receptor pair in human cancers, in which it regulates cell adhesion, extravasation, metastatic colonization, angiogenesis, and proliferation. All of these processes require activation of signaling pathways that include G proteins, phosphatidylinositol-3 kinase (PI3K), JAK kinases, Rho GTPases, and focal adhesion-associated proteins. We analyzed these pathways in a human melanoma cell line in response to CXCL12 stimulation, and found that PI3K{gamma} regulates tumor cell adhesion through mechanisms different from those involved in cell invasion. Our data indicate that, following CXCR4 activation after CXCL12 binding, the invasion and adhesion processes are regulated differently by distinct downstream events in these signaling cascades.

  18. Selective loss of gastrointestinal mast cells and impaired immunity in PI3K-deficient mice.

    PubMed

    Fukao, Taro; Yamada, Taketo; Tanabe, Masanobu; Terauchi, Yasuo; Ota, Takayuki; Takayama, Tetsuro; Asano, Tomoichiro; Takeuchi, Tsutomu; Kadowaki, Takashi; Hata Ji, Jun-ichi; Koyasu, Shigeo

    2002-03-01

    Mice that lack the p85alpha regulatory subunit of phosphatidylinositol-3 kinase (PI3K) are deficient in gastrointestinal and peritoneal mast cells but have dermal mast cells. Accordingly, these mice show impaired bacterial clearance in response to acute septic peritonitis and are highly susceptible to infection by the intestinal nematode Strongyloides venezuelensis. Systemic anaphylactic shock responses, however, are intact. We found that although reconstitution of PI3Kminus sign/minus sign mice with bone marrow--derived mast cells (BMMCs) restored anti-bacterial immunity, only T helper type 2 (TH2)-conditioned BMMCs, not "standard" BMMCs, were able to restore anti-nematode immunity. This finding highlights the importance of the TH2 response in the control of nematode infection. Thus, PI3K likely plays an essential role in host immune responses by regulating both the development and induction of mast cells.

  19. Clinical development of phosphatidylinositol-3 kinase pathway inhibitors.

    PubMed

    Arteaga, Carlos L

    2010-01-01

    The PI3K pathway is the most commonly altered in human cancer. Several recent phase I studies with therapeutic inhibitors of this pathway have shown that pharmacological inhibition of PI3K in humans is feasible and overall well tolerated. Furthermore, there has already been clinical evidence of anti-tumor activity in patients with advanced cancer. The intensity and duration of PI3K inhibition required for an antitumor effect and the optimal pharmacodynamic biomarker(s) of pathway inactivation remain to be established. Preclinical and early clinical data support focusing on trials with PI3K inhibitors that are at a minimum enriched with patients with alterations in this signaling pathway. These inhibitors are likely to be more effective in combination with established and other novel molecular therapies.

  20. Phosphatidylinositol-3-kinase p110γ Contributes to Bile Salt-Induced Apoptosis in Primary Rat Hepatocytes and Human Hepatoma Cells

    PubMed Central

    Hohenester, Simon; Gates, Anna; Wimmer, Ralf; Beuers, Ulrich; Anwer, M. Sawkat; Rust, Christian; Webster, Cynthia R.L.

    2010-01-01

    Background & Aims Glycochenodeoxycholate (GCDC) and taurolithocholate (TLC) are hepatotoxic and cholestatic bile salts, whereas tauroursodeoxycholate (TUDC) is cytoprotective and anticholestatic. Yet they all act, in part, through phosphatidylinositol-3-kinase(PI3K)-dependent mechanisms (“PI3K-paradox”). Hepatocytes express three catalytic PI3K Class I isoforms (p110α/β/γ), specific functions of which, in liver, are unclear. In other cell types, p110γ is associated with detrimental effects. To shed light on the PI3K enigma, we determined whether hydrophobic and hydrophilic bile salts differentially activate distinct p110 isoforms in hepatocytes, and whether p110γ mediates bile salt-induced hepatocyte cell death. Methods Isoform-specific PI3K activity assays were established to determine isoform activation by bile salts in rat hepatocytes. Activation of Akt and JNK were determined by immunoblotting. Following stimulation with hydrophobic bile salts, hepatocellular apoptosis was determined morphologically after Hoechst staining and by analysis of caspase-3/-7 activity or caspase-3 cleavage. Activity or expression of PI3K p110γ was inhibited pharmacologically (AS604850) or by knockdown using specific siRNA. Results All bile salts tested activated p110β, while p110α was activated by TUDC and GCDC. Intriguingly, only hydrophobic bile salts activated p110γ. Inhibition of p110γ attenuated GCDC-induced Akt- and JNK-activation, but did not alter TUDC- or cAMP-induced Akt signaling in rat hepatocytes. Inhibition or knockdown of p110γ markedly attenuated hydrophobic bile salt-induced apoptosis in rat hepatocytes and human hepatoma cell lines but did not alter Fas-, tumor necrosis factor α- and etoposide-induced apoptosis. Depletion of Ca++ prevented GCDC-induced toxicity in rat hepatocytes but did not affect GCDC-induced Akt- and JNK-activation. Conclusions PI3K p110γ is activated by hydrophobic, but not hydrophilic bile salts. Bile salt-induced hepatocyte

  1. Phosphatidylinositol-3-kinase p110γ contributes to bile salt-induced apoptosis in primary rat hepatocytes and human hepatoma cells.

    PubMed

    Hohenester, Simon; Gates, Anna; Wimmer, Ralf; Beuers, Ulrich; Anwer, M Sawkat; Rust, Christian; Webster, Cynthia R L

    2010-11-01

    Glycochenodeoxycholate (GCDC) and taurolithocholate (TLC) are hepatotoxic and cholestatic bile salts, whereas tauroursodeoxycholate (TUDC) is cytoprotective and anticholestatic. Yet they all act, in part, through phosphatidylinositol-3-kinase(PI3K)-dependent mechanisms ("PI3K-paradox"). Hepatocytes express three catalytic PI3K Class I isoforms (p110α/β/γ), specific functions of which, in liver, are unclear. In other cell types, p110γ is associated with detrimental effects. To shed light on the PI3K enigma, we determined whether hydrophobic and hydrophilic bile salts differentially activate distinct p110 isoforms in hepatocytes, and whether p110γ mediates bile salt-induced hepatocyte cell death. Isoform-specific PI3K activity assays were established to determine isoform activation by bile salts in rat hepatocytes. Activation of Akt and JNK was determined by immunoblotting. Following stimulation with hydrophobic bile salts, hepatocellular apoptosis was determined morphologically after Hoechst staining and by analysis of caspase-3/-7 activity or caspase-3 cleavage. Activity or expression of PI3K p110γ was inhibited pharmacologically (AS604850) or by knock-down using specific siRNA. All bile salts tested activated p110β, while p110α was activated by TUDC and GCDC. Intriguingly, only hydrophobic bile salts activated p110γ. Inhibition of p110γ attenuated GCDC-induced Akt- and JNK-activation, but did not alter TUDC- or cAMP-induced Akt-signaling in rat hepatocytes. Inhibition or knock-down of p110γ markedly attenuated hydrophobic bile salt-induced apoptosis in rat hepatocytes and human hepatoma cell lines but did not alter Fas-, tumor necrosis factor α- and etoposide-induced apoptosis. Depletion of Ca(++) prevented GCDC-induced toxicity in rat hepatocytes but did not affect GCDC-induced Akt- and JNK-activation. PI3K p110γ is activated by hydrophobic, but not hydrophilic bile salts. Bile salt-induced hepatocyte apoptosis is partly mediated via a PI3K p110

  2. Distinct regulation of autophagic activity by Atg14L and Rubicon associated with Beclin 1-phosphatidylinositol-3-kinase complex.

    PubMed

    Zhong, Yun; Wang, Qing Jun; Li, Xianting; Yan, Ying; Backer, Jonathan M; Chait, Brian T; Heintz, Nathaniel; Yue, Zhenyu

    2009-04-01

    Beclin 1, a mammalian autophagy protein that has been implicated in development, tumour suppression, neurodegeneration and cell death, exists in a complex with Vps34, the class III phosphatidylinositol-3-kinase (PI(3)K) that mediates multiple vesicle-trafficking processes including endocytosis and autophagy. However, the precise role of the Beclin 1-Vps34 complex in autophagy regulation remains to be elucidated. Combining mouse genetics and biochemistry, we have identified a large in vivo Beclin 1 complex containing the known proteins Vps34, p150/Vps15 and UVRAG, as well as two newly identified proteins, Atg14L (yeast Atg14-like) and Rubicon (RUN domain and cysteine-rich domain containing, Beclin 1-interacting protein). Characterization of the new proteins revealed that Atg14L enhances Vps34 lipid kinase activity and upregulates autophagy, whereas Rubicon reduces Vps34 activity and downregulates autophagy. We show that Beclin 1 and Atg14L synergistically promote the formation of double-membraned organelles that are associated with Atg5 and Atg12, whereas forced expression of Rubicon results in aberrant late endosomal/lysosomal structures and impaired autophagosome maturation. We hypothesize that by forming distinct protein complexes, Beclin 1 and its binding proteins orchestrate the precise function of the class III PI(3)K in regulating autophagy at multiple steps.

  3. Epidermal growth factor stimulates Rac activation through Src and phosphatidylinositol 3-kinase to promote colonic epithelial cell migration.

    PubMed

    Dise, Rebecca S; Frey, Mark R; Whitehead, Robert H; Polk, D Brent

    2008-01-01

    Regulated intestinal epithelial cell migration plays a key role in wound healing and maintenance of a healthy gastrointestinal tract. Epidermal growth factor (EGF) stimulates cell migration and wound closure in intestinal epithelial cells through incompletely understood mechanisms. In this study we investigated the role of the small GTPase Rac in EGF-induced cell migration using an in vitro wound-healing assay. In mouse colonic epithelial (MCE) cell lines, EGF-stimulated wound closure was accompanied by a doubling of the number of cells containing lamellipodial extensions at the wound margin, increased Rac membrane translocation in cells at the wound margin, and rapid Rac activation. Either Rac1 small interfering (si)RNA or a Rac1 inhibitor completely blocked EGF-stimulated wound closure. Whereas EGF failed to activate Rac in colon cells from EGF receptor (EGFR) knockout mice, stable expression of wild-type EGFR restored EGF-stimulated Rac activation and migration. Pharmacological inhibition of either phosphatidylinositol 3-kinase (PI3K) or Src family kinases reduced EGF-stimulated Rac activation. Cotreatment of cells with both inhibitors completely blocked EGF-stimulated Rac activation and localization to the leading edge of cells and lamellipodial extension. Our results present a novel mechanism by which the PI3K and Src signaling cascades cooperate to activate Rac and promote intestinal epithelial cell migration downstream of EGFR.

  4. The role of the PI3K-Akt signal transduction pathway in Autographa californica multiple nucleopolyhedrovirus infection of Spodoptera frugiperda cells

    SciTech Connect

    Xiao Wei; Yang Yi; Weng Qingbei; Lin Tiehao; Yuan Meijin; Yang Kai; Pang Yi

    2009-08-15

    Many viruses activate the phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway, thereby modulating diverse downstream signaling pathways associated with antiapoptosis, proliferation, cell cycling, protein synthesis and glucose metabolism, in order to augment their replication. To date, the role of the PI3K-Akt pathway in Baculovirus replication has not been defined. In the present study, we demonstrate that infection of Sf9 cells with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) elevated cellular Akt phosphorylation at 1 h post-infection. The maximum Akt phosphorylation occurred at 6 h post-infection and remained unchanged until 18 h post-infection. The PI3K-specific inhibitor, LY294002, suppressed Akt phosphorylation in a dose-dependent manner, suggesting that AcMNPV-induced Akt phosphorylation is PI3K-dependent. The inhibition of PI3K-Akt activation by LY294002 significantly reduced the viral yield, including a reduction in budded viruses and occlusion bodies. The virus production was reduced only when the inhibitor was added within 24 h of infection, implying that activation of PI3K occurred early in infection. Correspondingly, both viral DNA replication and late (VP39) and very late (POLH) viral protein expression were impaired by LY294002 treatment; LY294002 had no effect on immediate-early (IE1) and early-late (GP64) protein expression. These results demonstrate that the PI3K-Akt pathway is required for efficient Baculovirus replication.

  5. Platelet-derived growth factor-dependent cellular transformation requires either phospholipase Cgamma or phosphatidylinositol 3 kinase.

    PubMed

    DeMali, K A; Whiteford, C C; Ulug, E T; Kazlauskas, A

    1997-04-04

    Although it has been well established that constitutive activation of receptor tyrosine kinases leads to cellular transformation, the signal relay pathways involved have not been systematically investigated. In this study we used a panel of platelet-derived growth factor (PDGF) beta receptor mutants (beta-PDGFR), which selectively activate various signal relay enzymes to define which signaling pathways are required for PDGF-dependent growth of cells in soft agar. The host cell line for these studies was Ph cells, a 3T3-like cell that expresses normal levels of the beta-PDGFR but no PDGF-alpha receptor (alpha-PDGFR). Hence, this cell system can be used to study signaling of mutant alphaPDGFRs or alpha/beta chimeras. We constructed chimeric receptors containing the alphaPDGFR extracellular domain and the betaPDGFR cytoplasmic domain harboring various phosphorylation site mutations. The mutants were expressed in Ph cells, and their ability to drive PDGF-dependent cellular transformation (growth in soft agar) was assayed. Cells infected with an empty expression vector failed to grow in soft agar, whereas introduction of the chimera with a wild-type beta-PDGFR cytoplasmic domain gave rise to a large number of colonies. In contrast, the N2F5 chimera, in which the binding sites for phospholipase Cgamma (PLC-gamma), RasGTPase-activating protein, phosphatidylinositol 3 kinase (PI3K), and SHP-2 were eliminated, failed to trigger proliferation. Restoring the binding sites for RasGTPase-activating protein or SHP-2 did not rescue the PDGF-dependent response. In contrast, receptors capable of associating with either PLC-gamma or PI3K relayed a growth signal that was comparable to wild-type receptors in the soft agar growth assay. These findings indicate that the PDGF receptor activates multiple signaling pathways that lead to cellular transformation, and that either PI3K or PLC-gamma are key initiators of such signal relay cascades.

  6. Liver Dysfunction and Phosphatidylinositol-3-Kinase Signalling in Early Sepsis: Experimental Studies in Rodent Models of Peritonitis

    PubMed Central

    Westermann, Martin; Lambeck, Sandro; Lupp, Amelie; Rudiger, Alain; Dyson, Alex; Carré, Jane E.; Kortgen, Andreas; Krafft, Christoph; Popp, Jürgen; Sponholz, Christoph; Fuhrmann, Valentin; Hilger, Ingrid; Claus, Ralf A.; Riedemann, Niels C.; Wetzker, Reinhard; Singer, Mervyn; Trauner, Michael; Bauer, Michael

    2012-01-01

    Background Hepatic dysfunction and jaundice are traditionally viewed as late features of sepsis and portend poor outcomes. We hypothesized that changes in liver function occur early in the onset of sepsis, yet pass undetected by standard laboratory tests. Methods and Findings In a long-term rat model of faecal peritonitis, biotransformation and hepatobiliary transport were impaired, depending on subsequent disease severity, as early as 6 h after peritoneal contamination. Phosphatidylinositol-3-kinase (PI3K) signalling was simultaneously induced at this time point. At 15 h there was hepatocellular accumulation of bilirubin, bile acids, and xenobiotics, with disturbed bile acid conjugation and drug metabolism. Cholestasis was preceded by disruption of the bile acid and organic anion transport machinery at the canalicular pole. Inhibitors of PI3K partially prevented cytokine-induced loss of villi in cultured HepG2 cells. Notably, mice lacking the PI3Kγ gene were protected against cholestasis and impaired bile acid conjugation. This was partially confirmed by an increase in plasma bile acids (e.g., chenodeoxycholic acid [CDCA] and taurodeoxycholic acid [TDCA]) observed in 48 patients on the day severe sepsis was diagnosed; unlike bilirubin (area under the receiver-operating curve: 0.59), these bile acids predicted 28-d mortality with high sensitivity and specificity (area under the receiver-operating curve: CDCA: 0.77; TDCA: 0.72; CDCA+TDCA: 0.87). Conclusions Liver dysfunction is an early and commonplace event in the rat model of sepsis studied here; PI3K signalling seems to play a crucial role. All aspects of hepatic biotransformation are affected, with severity relating to subsequent prognosis. Detected changes significantly precede conventional markers and are reflected by early alterations in plasma bile acids. These observations carry important implications for the diagnosis of liver dysfunction and pharmacotherapy in the critically ill. Further clinical work is

  7. Promotion of melanoma cell invasion and tumor metastasis by microcystin-LR via phosphatidylinositol 3-kinase/AKT pathway.

    PubMed

    Xu, Pengfei; Zhang, Xu-Xiang; Miao, Chen; Fu, Ziyi; Li, Zhengrong; Zhang, Gen; Zheng, Maqing; Liu, Yuefang; Yang, Liuyan; Wang, Ting

    2013-08-06

    Recently, we have indicated that microcystin-LR, a cyanobacterial toxin produced in eutrophic lakes or reservoirs, can increase invasive ability of melanoma MDA-MB-435 cells; however, the stimulatory effect needs identification by in vivo experiment and the related molecular mechanism is poorly understood. In this study, in vitro and in vivo experiments were conducted to investigate the effect of microcystin-LR on invasion and metastasis of human melanoma cells, and the underlying molecular mechanism was also explored. MDA-MB-435 xenograft model assay showed that oral administration of nude mice with microcystin-LR at 0.001-0.1 mg/kg/d posed no significant effect on tumor weight. Histological examination demonstrated that microcystin-LR could promote lung metastasis, which is confirmed by Matrigel chamber assay suggesting that microcystin-LR treatment at 25 nM can increase the invasiveness of MDA-MB-435 cells. In vitro and in vivo experiments consistently showed that microcystin-LR exposure increased mRNA and protein levels of matrix metalloproteinases (MMP-2/-9) by activating phosphatidylinositol 3-kinase (PI3-K)/AKT. Additionally, microcystin-LR treatment at low doses (≤25 nM) decreased lipid phosphatase PTEN expression, and the microcystin-induced invasiveness enhancement and MMP-2/-9 overexpression were reversed by the PI3-K/AKT chemical inhibitor LY294002 and AKT siRNA, indicating that microcystin-LR promotes invasion and metastasis of MDA-MB-435 cells via the PI3-K/AKT pathway.

  8. Phosphatidylinositol 3-Kinase Mediates β-Catenin Dysfunction of Airway Epithelium in a Toluene Diisocyanate-Induced Murine Asthma Model.

    PubMed

    Yao, Lihong; Zhao, Haijin; Tang, Haixiong; Song, Jiafu; Dong, Hangming; Zou, Fei; Cai, Shaoxi

    2015-09-01

    Cell-cell junctions are critical for the maintenance of cellular as well as tissue polarity and integrity. Yet the role of phosphatidylinositol 3-kinase (PI3K) in dysregulation of airway epithelial adherens junctions in toluene diisocyanate (TDI)-induced asthma has not been addressed. Male BALB/c mice were first dermally sensitized and then challenged with TDI by means of compressed air nebulization. The mice were treated intratracheally with PI3K inhibitor LY294002. Levels of phospho-Akt in airway epithelium and whole lung tissues were markedly increased in TDI group compared with control mice, which decreased after administration of LY294002. The dilated intercellular spaces of airway epithelium induced by TDI were partially recovered by LY294002. Both the protein expression and distribution of adherens junction proteins E-cadherin and β-catenin were altered by TDI. Treatment with LY294002 rescued the distribution of E-cadherin and β-catenin at cell-cell membranes, restored total β-catenin pool, but had no effect on protein level of E-cadherin. At the same time, LY294002 also inhibited phosphorylation of ERK, glycogen synthase kinase3β and tyrosine 654 of β-catenin induced by TDI. In summary, our results showed that the PI3K pathway mediates β-catenin dysregulation in a TDI-induced murine asthma model, which may be associated with increased tyrosine phosphorylation of β-catenin. © The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  9. Randomized phase 1 study of the phosphatidylinositol 3-kinase δ inhibitor idelalisib in patients with allergic rhinitis.

    PubMed

    Horak, Friedrich; Puri, Kamal D; Steiner, Bart H; Holes, Leanne; Xing, Guan; Zieglmayer, Petra; Zieglmayer, René; Lemell, Patrick; Yu, Albert

    2016-06-01

    Phosphatidylinositol 3-kinase p110δ isoform (PI3K p110δ) activity is essential for mast cell activation, suggesting that inhibition of PI3K p110δ might be useful in treating allergic diseases. We sought to determine the effect of the PI3K p110δ-selective inhibitor idelalisib on allergic responses. This phase 1 randomized, double-blind, placebo-controlled, 2-period crossover study was conducted with the Vienna Challenge Chamber. Grass pollen-induced allergic symptoms were documented during screening. Eligible subjects received idelalisib (100 mg twice daily) or placebo for 7 days, with allergen challenge on day 7. After a 2-week washout period, subjects received the alternate treatment and repeated allergen challenge. Study measures included safety, nasal and nonnasal symptoms, nasal airflow, nasal secretions, basophil activation, and plasma cytokine levels. Forty-one patients with allergic rhinitis received idelalisib/placebo (n = 21) or placebo/idelalisib (n = 20). Idelalisib treatment was well tolerated. Mean total nasal symptom scores were lower during the combined idelalisib treatment periods compared with placebo (treatment difference [idelalisib - placebo], -1.78; 95% CI, -2.53 to -1.03; P < .001). Statistically significant differences were also observed for the combined treatment periods for total symptom scores, nasal airflow, nasal secretion weight, and nasal congestion scores. The percentage of ex vivo-activated basophils (CD63(+)/CCR3(+) cells; after stimulation with grass pollen) was substantially lower for idelalisib-treated compared with placebo-treated subjects. Plasma CCL17 and CCL22 levels were reduced after idelalisib treatment. Idelalisib treatment was well tolerated in patients with allergic rhinitis and appears to reduce allergic responses clinically and immunologically after an environmental allergen challenge. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  10. PI3K-GSK3 signalling regulates mammalian axon regeneration by inducing the expression of Smad1

    NASA Astrophysics Data System (ADS)

    Saijilafu; Hur, Eun-Mi; Liu, Chang-Mei; Jiao, Zhongxian; Xu, Wen-Lin; Zhou, Feng-Quan

    2013-10-01

    In contrast to neurons in the central nervous system, mature neurons in the mammalian peripheral nervous system (PNS) can regenerate axons after injury, in part, by enhancing intrinsic growth competence. However, the signalling pathways that enhance the growth potential and induce spontaneous axon regeneration remain poorly understood. Here we reveal that phosphatidylinositol 3-kinase (PI3K) signalling is activated in response to peripheral axotomy and that PI3K pathway is required for sensory axon regeneration. Moreover, we show that glycogen synthase kinase 3 (GSK3), rather than mammalian target of rapamycin, mediates PI3K-dependent augmentation of the growth potential in the PNS. Furthermore, we show that PI3K-GSK3 signal is conveyed by the induction of a transcription factor Smad1 and that acute depletion of Smad1 in adult mice prevents axon regeneration in vivo. Together, these results suggest PI3K-GSK3-Smad1 signalling as a central module for promoting sensory axon regeneration in the mammalian nervous system.

  11. Effects of AFP-activated PI3K/Akt signaling pathway on cell proliferation of liver cancer.

    PubMed

    Zheng, Lu; Gong, Wei; Liang, Ping; Huang, XiaoBing; You, Nan; Han, Ke Qiang; Li, Yu Ming; Li, Jing

    2014-05-01

    This study aims to investigate effects of alpha-fetoprotein (AFP)-activated phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway on hepatocellular carcinoma cell proliferation. Active cirrhosis patients after hepatitis B infection (n = 20) and viral hepatitis patients with hepatocellular carcinoma (HCC) (n = 20) were selected as the subjects of the present study. Another 20 healthy subjects were selected as the control group. The serum AFP expression and liver tissue PI3K and Akt gene mRNA expression were detected. The hepatoma cell model HepG2 which had a stable expression of AFP gene was used. Real-time quantitative PCR and Western blot and other methods were used to analyze the intracellular PI3K and Akt protein levels. Compared with control group and cirrhosis group, the serum AFP levels in HCC group significantly increased, and the tissue PI3K and Akt mRNA expression also significantly increased. HepG2 cells were intervened using AFP, in which the PIK and Akt protein expression significantly increased. After intervention by use of AFP monoclonal antibodies or LY294002 inhibitor, the PIK and Akt protein expression in HepG2 cell was significantly decreased (P < 0.05). AFP can promote the proliferation of hepatoma cells via activation of PI3K/Akt signaling pathway.

  12. Programmed Death-1 Inhibition of Phosphatidylinositol 3-Kinase/AKT/Mechanistic Target of Rapamycin Signaling Impairs Sarcoidosis CD4(+) T Cell Proliferation.

    PubMed

    Celada, Lindsay J; Rotsinger, Joseph E; Young, Anjuli; Shaginurova, Guzel; Shelton, Debresha; Hawkins, Charlene; Drake, Wonder P

    2017-01-01

    Patients with progressive sarcoidosis exhibit increased expression of programmed death-1 (PD-1) receptor on their CD4(+) T cells. Up-regulation of this marker of T cell exhaustion is associated with a reduction in the proliferative response to T cell receptor (TCR) stimulation, a defect that is reversed by PD-1 pathway blockade. Genome-wide association studies and microarray analyses have correlated signaling downstream from the TCR with sarcoidosis disease severity, but the mechanism is not yet known. Reduced phosphatidylinositol 3-kinase (PI3K)/AKT expression inhibits proliferation by inhibiting cell cycle progression. To test the hypothesis that PD-1 expression attenuates TCR-dependent activation of PI3K/AKT activity in progressive systemic sarcoidosis, we analyzed PI3K/AKT/mechanistic target of rapamycin (mTOR) expression at baseline and after PD-1 pathway blockade in CD4(+) T cells isolated from patients with sarcoidosis and healthy control subjects. We confirmed an increased percentage of PD-1(+) CD4(+) T cells and reduced proliferative capacity in patients with sarcoidosis compared with healthy control subjects (P < 0.001). There was a negative correlation with PD-1 expression and proliferative capacity (r = -0.70, P < 0.001). Expression of key mediators of cell cycle progression, including PI3K and AKT, were significantly decreased. Gene and protein expression levels reverted to healthy control levels after PD-1 pathway blockade. Reduction in sarcoidosis CD4(+) T cell proliferative capacity is secondary to altered expression of key mediators of cell cycle progression, including the PI3K/AKT/mTOR pathway, via PD-1 up-regulation. This supports the concept that PD-1 up-regulation drives the immunologic deficits associated with sarcoidosis severity by inducing signaling aberrancies in key mediators of cell cycle progression.

  13. A Phosphatidylinositol 3-Kinase/Protein Kinase B-independent Activation of Mammalian Target of Rapamycin Signaling Is Sufficient to Induce Skeletal Muscle Hypertrophy

    PubMed Central

    Goodman, Craig A.; Miu, Man Hing; Frey, John W.; Mabrey, Danielle M.; Lincoln, Hannah C.; Ge, Yejing; Chen, Jie

    2010-01-01

    It has been widely proposed that signaling by mammalian target of rapamycin (mTOR) is both necessary and sufficient for the induction of skeletal muscle hypertrophy. Evidence for this hypothesis is largely based on studies that used stimuli that activate mTOR via a phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB)-dependent mechanism. However, the stimulation of signaling by PI3K/PKB also can activate several mTOR-independent growth-promoting events; thus, it is not clear whether signaling by mTOR is permissive, or sufficient, for the induction of hypertrophy. Furthermore, the presumed role of mTOR in hypertrophy is derived from studies that used rapamycin to inhibit mTOR; yet, there is very little direct evidence that mTOR is the rapamycin-sensitive element that confers the hypertrophic response. In this study, we determined that, in skeletal muscle, overexpression of Rheb stimulates a PI3K/PKB-independent activation of mTOR signaling, and this is sufficient for the induction of a rapamycin-sensitive hypertrophic response. Transgenic mice with muscle specific expression of various mTOR mutants also were used to demonstrate that mTOR is the rapamycin-sensitive element that conferred the hypertrophic response and that the kinase activity of mTOR is necessary for this event. Combined, these results provide direct genetic evidence that a PI3K/PKB-independent activation of mTOR signaling is sufficient to induce hypertrophy. In summary, overexpression of Rheb activates mTOR signaling via a PI3K/PKB-independent mechanism and is sufficient to induce skeletal muscle hypertrophy. The hypertrophic effects of Rheb are driven through a rapamycin-sensitive (RS) mechanism, mTOR is the RS element that confers the hypertrophy, and the kinase activity of mTOR is necessary for this event. PMID:20668162

  14. A phosphatidylinositol 3-kinase/protein kinase B-independent activation of mammalian target of rapamycin signaling is sufficient to induce skeletal muscle hypertrophy.

    PubMed

    Goodman, Craig A; Miu, Man Hing; Frey, John W; Mabrey, Danielle M; Lincoln, Hannah C; Ge, Yejing; Chen, Jie; Hornberger, Troy A

    2010-09-15

    It has been widely proposed that signaling by mammalian target of rapamycin (mTOR) is both necessary and sufficient for the induction of skeletal muscle hypertrophy. Evidence for this hypothesis is largely based on studies that used stimuli that activate mTOR via a phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB)-dependent mechanism. However, the stimulation of signaling by PI3K/PKB also can activate several mTOR-independent growth-promoting events; thus, it is not clear whether signaling by mTOR is permissive, or sufficient, for the induction of hypertrophy. Furthermore, the presumed role of mTOR in hypertrophy is derived from studies that used rapamycin to inhibit mTOR; yet, there is very little direct evidence that mTOR is the rapamycin-sensitive element that confers the hypertrophic response. In this study, we determined that, in skeletal muscle, overexpression of Rheb stimulates a PI3K/PKB-independent activation of mTOR signaling, and this is sufficient for the induction of a rapamycin-sensitive hypertrophic response. Transgenic mice with muscle specific expression of various mTOR mutants also were used to demonstrate that mTOR is the rapamycin-sensitive element that conferred the hypertrophic response and that the kinase activity of mTOR is necessary for this event. Combined, these results provide direct genetic evidence that a PI3K/PKB-independent activation of mTOR signaling is sufficient to induce hypertrophy. In summary, overexpression of Rheb activates mTOR signaling via a PI3K/PKB-independent mechanism and is sufficient to induce skeletal muscle hypertrophy. The hypertrophic effects of Rheb are driven through a rapamycin-sensitive (RS) mechanism, mTOR is the RS element that confers the hypertrophy, and the kinase activity of mTOR is necessary for this event.

  15. The Neuron-Specific Rai (ShcC) Adaptor Protein Inhibits Apoptosis by Coupling Ret to the Phosphatidylinositol 3-Kinase/Akt Signaling Pathway

    PubMed Central

    Pelicci, Giuliana; Troglio, Flavia; Bodini, Alessandra; Melillo, Rosa Marina; Pettirossi, Valentina; Coda, Laura; De Giuseppe, Antonio; Santoro, Massimo; Pelicci, Pier Giuseppe

    2002-01-01

    Rai is a recently identified member of the family of Shc-like proteins, which are cytoplasmic signal transducers characterized by the unique PTB-CH1-SH2 modular organization. Rai expression is restricted to neuronal cells and regulates in vivo the number of postmitotic sympathetic neurons. We report here that Rai is not a common substrate of receptor tyrosine kinases under physiological conditions and that among the analyzed receptors (Ret, epidermal growth factor receptor, and TrkA) it is activated specifically by Ret. Overexpression of Rai in neuronal cell lines promoted survival by reducing apoptosis both under conditions of limited availability of the Ret ligand glial cell line-derived neurotrophic factor (GDNF) and in the absence of Ret activation. Overexpressed Rai resulted in the potentiation of the Ret-dependent activation of phosphatidylinositol 3-kinase (PI3K) and Akt. Notably, increased Akt phosphorylation and PI3K activity were also found under basal conditions, e.g., in serum-starved neuronal cells. Phosphorylated and hypophosphorylated Rai proteins form a constitutive complex with the p85 subunit of PI3K: upon Ret triggering, the Rai-PI3K complex is recruited to the tyrosine-phosphorylated Ret receptor through the binding of the Rai PTB domain to tyrosine 1062 of Ret. In neurons treated with low concentrations of GDNF, the prosurvival effect of Rai depends on Rai phosphorylation and Ret activation. In the absence of Ret activation, the prosurvival effect of Rai is, instead, phosphorylation independent. Finally, we showed that overexpression of Rai, at variance with Shc, had no effects on the early peak of mitogen-activated protein kinase (MAPK) activation, whereas it increased its activation at later time points. Phosphorylated Rai, however, was not found in complexes with Grb2. We propose that Rai potentiates the MAPK and PI3K signaling pathways and regulates Ret-dependent and -independent survival signals. PMID:12242309

  16. Molecular targets for cancer therapy in the PI3K/AKT/mTOR pathway.

    PubMed

    Polivka, Jiri; Janku, Filip

    2014-05-01

    Aberrations in various cellular signaling pathways are instrumental in regulating cellular metabolism, tumor development, growth, proliferation, metastasis and cytoskeletal reorganization. The fundamental cellular signaling cascade involved in these processes, the phosphatidylinositol 3-kinase/protein kinase-B/mammalian target of rapamycin (PI3K/AKT/mTOR), closely related to the mitogen-activated protein kinase (MAPK) pathway, is a crucial and intensively explored intracellular signaling pathway in tumorigenesis. Various activating mutations in oncogenes together with the inactivation of tumor suppressor genes are found in diverse malignancies across almost all members of the pathway. Substantial progress in uncovering PI3K/AKT/mTOR alterations and their roles in tumorigenesis has enabled the development of novel targeted molecules with potential for developing efficacious anticancer treatment. Two approved anticancer drugs, everolimus and temsirolimus, exemplify targeted inhibition of PI3K/AKT/mTOR in the clinic and many others are in preclinical development as well as being tested in early clinical trials for many different types of cancer. This review focuses on targeted PI3K/AKT/mTOR signaling from the perspective of novel molecular targets for cancer therapy found in key pathway members and their corresponding experimental therapeutic agents. Various aberrant prognostic and predictive biomarkers are also discussed and examples are given. Novel approaches to PI3K/AKT/mTOR pathway inhibition together with a better understanding of prognostic and predictive markers have the potential to significantly improve the future care of cancer patients in the current era of personalized cancer medicine.

  17. PI-103 and Quercetin Attenuate PI3K-AKT Signaling Pathway in T- Cell Lymphoma Exposed to Hydrogen Peroxide.

    PubMed

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2016-01-01

    Phosphatidylinositol 3 kinase-protein kinase B (PI3K-AKT) pathway has been considered as major drug target site due to its frequent activation in cancer. AKT regulates the activity of various targets to promote tumorigenesis and metastasis. Accumulation of reactive oxygen species (ROS) has been linked to oxidative stress and regulation of signaling pathways for metabolic adaptation of tumor microenvironment. Hydrogen peroxide (H2O2) in this context is used as ROS source for oxidative stress preconditioning. Antioxidants are commonly considered to be beneficial to reduce detrimental effects of ROS and are recommended as dietary supplements. Quercetin, a ubiquitous bioactive flavonoid is a dietary component which has attracted much of interest due to its potential health-promoting effects. Present study is aimed to analyze PI3K-AKT signaling pathway in H2O2 exposed Dalton's lymphoma ascite (DLA) cells. Further, regulation of PI3K-AKT pathway by quercetin as well as PI-103, an inhibitor of PI3K was analyzed. Exposure of H2O2 (1mM H2O2 for 30min) to DLA cells caused ROS accumulation and resulted in increased phosphorylation of PI3K and downstream proteins PDK1 and AKT (Ser-473 and Thr-308), cell survival factors BAD and ERK1/2, as well as TNFR1. However, level of tumor suppressor PTEN was declined. Both PI-103 & quercetin suppressed the enhanced level of ROS and significantly down-regulated phosphorylation of AKT, PDK1, BAD and level of TNFR1 as well as increased the level of PTEN in H2O2 induced lymphoma cells. The overall result suggests that quercetin and PI3K inhibitor PI-103 attenuate PI3K-AKT pathway in a similar mechanism.

  18. Increased persistent sodium current due to decreased PI3K signaling contributes to QT prolongation in the diabetic heart.

    PubMed

    Lu, Zhongju; Jiang, Ya-Ping; Wu, Chia-Yen C; Ballou, Lisa M; Liu, Shengnan; Carpenter, Eileen S; Rosen, Michael R; Cohen, Ira S; Lin, Richard Z

    2013-12-01

    Diabetes is an independent risk factor for sudden cardiac death and ventricular arrhythmia complications of acute coronary syndrome. Prolongation of the QT interval on the electrocardiogram is also a risk factor for arrhythmias and sudden death, and the increased prevalence of QT prolongation is an independent risk factor for cardiovascular death in diabetic patients. The pathophysiological mechanisms responsible for this lethal complication are poorly understood. Diabetes is associated with a reduction in phosphoinositide 3-kinase (PI3K) signaling, which regulates the action potential duration (APD) of individual myocytes and thus the QT interval by altering multiple ion currents, including the persistent sodium current INaP. Here, we report a mechanism for diabetes-induced QT prolongation that involves an increase in INaP caused by defective PI3K signaling. Cardiac myocytes of mice with type 1 or type 2 diabetes exhibited an increase in APD that was reversed by expression of constitutively active PI3K or intracellular infusion of phosphatidylinositol 3,4,5-trisphosphate (PIP3), the second messenger produced by PI3K. The diabetic myocytes also showed an increase in INaP that was reversed by activated PI3K or PIP3. The increases in APD and INaP in myocytes translated into QT interval prolongation for both types of diabetic mice. The long QT interval of type 1 diabetic hearts was shortened by insulin treatment ex vivo, and this effect was blocked by a PI3K inhibitor. Treatment of both types of diabetic mouse hearts with an INaP blocker also shortened the QT interval. These results indicate that downregulation of cardiac PI3K signaling in diabetes prolongs the QT interval at least in part by causing an increase in INaP. This mechanism may explain why the diabetic population has an increased risk of life-threatening arrhythmias.

  19. Phosphatidylinositol 3-Kinase: A Link Between Inflammation and Pancreatic Cancer

    PubMed Central

    Birtolo, Chiara; Go, Vay Liang W.; Ptasznik, Andrzej; Eibl, Guido; Pandol, Stephen J.

    2016-01-01

    Even though a strong association between inflammation and cancer has been widely accepted, the underlying precise molecular mechanisms are still largely unknown. A complex signaling network between tumor and stromal cells is responsible for the infiltration of inflammatory cells into the cancer micro-environment. Tumor stromal cells such as pancreatic stellate cells (PSCs) and immune cells create a microenvironment that protects cancer cells through a complex interaction, ultimately facilitating their local proliferation and their migration to different sites. Furthermore, PSCs have multiple functions related to local immunity, angiogenesis, inflammation and fibrosis. Recently, many studies have shown that members of the phosphoinositol-3-phosphate kinase (PI3K) family are activated in tumor cells, PSCs and tumor infiltrating inflammatory cells to promote cancer growth. Pro-inflammatory cytokines and chemokines secreted by immune cells and fibroblasts within the tumor environment can activate the PI3K pathway both in cancer and inflammatory cells. In this review, we focus on the central role of the PI3K pathway in regulating the cross-talk between immune/stromal cells and cancer cells. Understanding the role of the PI3K pathway in the development of chronic pancreatitis and cancer is crucial for the discovery of novel and efficacious treatment options. PMID:26658038

  20. Positron emission tomographic monitoring of dual phosphatidylinositol-3-kinase and mTOR inhibition in anaplastic large cell lymphoma

    PubMed Central

    Graf, Nicolas; Li, Zhoulei; Herrmann, Ken; Weh, Daniel; Aichler, Michaela; Slawska, Jolanta; Walch, Axel; Peschel, Christian; Schwaiger, Markus; Buck, Andreas K; Dechow, Tobias; Keller, Ulrich

    2014-01-01

    Background Dual phosphatidylinositol-3-kinase (PI3K)/mammalian target of rapamycin (mTOR) inhibition offers an attractive therapeutic strategy in anaplastic large cell lymphoma depending on oncogenic nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) signaling. We tested the efficacy of a novel dual PI3K/mTOR inhibitor, NVP-BGT226 (BGT226), in two anaplastic large cell lymphoma cell lines in vitro and in vivo and performed an early response evaluation with positron emission tomography (PET) imaging using the standard tracer, 2-deoxy-2-[18F]fluoro-D-glucose (FDG) and the thymidine analog, 3′-deoxy-3′-[18F] fluorothymidine (FLT). Methods The biological effects of BGT226 were determined in vitro in the NPM-ALK positive cell lines SU-DHL-1 and Karpas299 by 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, propidium iodide staining, and biochemical analysis of PI3K and mTOR downstream signaling. FDG-PET and FLT-PET were performed in immunodeficient mice bearing either SU-DHL-1 or Karpas299 xenografts at baseline and 7 days after initiation of treatment with BGT226. Lymphomas were removed for immunohistochemical analysis of proliferation and apoptosis to correlate PET findings with in vivo treatment effects. Results SU-DHL-1 cells showed sensitivity to BGT226 in vitro, with cell cycle arrest in G0/G1 phase and an IC50 in the low nanomolar range, in contrast with Karpas299 cells, which were mainly resistant to BGT226. In vivo, both FDG-PET and FLT-PET discriminated sensitive from resistant lymphoma, as indicated by a significant reduction of tumor-to-background ratios on day 7 in treated SU-DHL-1 lymphoma-bearing animals compared with the control group, but not in animals with Karpas299 xenografts. Imaging results correlated with a marked decrease in the proliferation marker Ki67, and a slight increase in the apoptotic marker, cleaved caspase 3, as revealed by immunostaining of explanted lymphoma tissue. Conclusion Dual PI3K/mTOR inhibition using BGT

  1. Positron emission tomographic monitoring of dual phosphatidylinositol-3-kinase and mTOR inhibition in anaplastic large cell lymphoma.

    PubMed

    Graf, Nicolas; Li, Zhoulei; Herrmann, Ken; Weh, Daniel; Aichler, Michaela; Slawska, Jolanta; Walch, Axel; Peschel, Christian; Schwaiger, Markus; Buck, Andreas K; Dechow, Tobias; Keller, Ulrich

    2014-01-01

    Dual phosphatidylinositol-3-kinase (PI3K)/mammalian target of rapamycin (mTOR) inhibition offers an attractive therapeutic strategy in anaplastic large cell lymphoma depending on oncogenic nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) signaling. We tested the efficacy of a novel dual PI3K/mTOR inhibitor, NVP-BGT226 (BGT226), in two anaplastic large cell lymphoma cell lines in vitro and in vivo and performed an early response evaluation with positron emission tomography (PET) imaging using the standard tracer, 2-deoxy-2-[(18)F]fluoro-D-glucose (FDG) and the thymidine analog, 3'-deoxy-3'-[(18)F] fluorothymidine (FLT). The biological effects of BGT226 were determined in vitro in the NPM-ALK positive cell lines SU-DHL-1 and Karpas299 by 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, propidium iodide staining, and biochemical analysis of PI3K and mTOR downstream signaling. FDG-PET and FLT-PET were performed in immunodeficient mice bearing either SU-DHL-1 or Karpas299 xenografts at baseline and 7 days after initiation of treatment with BGT226. Lymphomas were removed for immunohistochemical analysis of proliferation and apoptosis to correlate PET findings with in vivo treatment effects. SU-DHL-1 cells showed sensitivity to BGT226 in vitro, with cell cycle arrest in G0/G1 phase and an IC50 in the low nanomolar range, in contrast with Karpas299 cells, which were mainly resistant to BGT226. In vivo, both FDG-PET and FLT-PET discriminated sensitive from resistant lymphoma, as indicated by a significant reduction of tumor-to-background ratios on day 7 in treated SU-DHL-1 lymphoma-bearing animals compared with the control group, but not in animals with Karpas299 xenografts. Imaging results correlated with a marked decrease in the proliferation marker Ki67, and a slight increase in the apoptotic marker, cleaved caspase 3, as revealed by immunostaining of explanted lymphoma tissue. Dual PI3K/mTOR inhibition using BGT226 is effective in ALK

  2. Interleukin-10-induced gene expression and suppressive function are selectively modulated by the PI3K-Akt-GSK3 pathway

    PubMed Central

    Antoniv, Taras T; Ivashkiv, Lionel B

    2011-01-01

    Interleukin-10 (IL-10) is an immunosuppressive cytokine that inhibits inflammatory gene expression. Phosphatidylinositol 3-kinase (PI3K) -mediated signalling regulates inflammatory responses and can induce IL-10 production, but a role for PI3K signalling in cellular responses to IL-10 is not known. In this study we investigated the involvement of the PI3K-Akt-GSK3 signalling pathway in IL-10-induced gene expression and IL-10-mediated suppression of Toll-like receptor-induced gene expression in primary human macrophages. A combination of loss and gain of function approaches using kinase inhibitors, expression of constitutively active Akt, and RNA interference in primary human macrophages showed that expression of a subset of IL-10-inducible genes was dependent on PI3K-Akt signalling. The effects of PI3K-Akt signalling on IL-10 responses were mediated at least in part by glycogen synthase kinase 3 (GSK3). In accordance with a functional role for PI3K pathways in contributing to the suppressive actions of IL-10, PI3K signalling augmented IL-10-mediated inhibition of lipopolysaccharide-induced IL-1, IL-8 and cyclo-oxygenase-2 expression. The PI3K signalling selectively modulated IL-10 responses, as it was not required for inhibition of tumour necrosis factor expression or for induction of certain IL-10-inducible genes such as SOCS3. These findings identify a new mechanism by which PI3K-mediated signalling can suppress inflammation by regulating IL-10-mediated gene induction and anti-inflammatory function. PMID:21255011

  3. Inhibition of class IA PI3K enzymes in non-small cell lung cancer cells uncovers functional compensation among isoforms.

    PubMed

    Stamatkin, Christopher; Ratermann, Kelley L; Overley, Colleen W; Black, Esther P

    2015-01-01

    Deregulation of the phosphatidylinositol 3-kinase (PI3K) pathway is central to many human malignancies while normal cell proliferation requires pathway functionality. Although inhibitors of the PI3K pathway are in clinical trials or approved for therapy, an understanding of the functional activities of pathway members in specific malignancies is needed. In lung cancers, the PI3K pathway is often aberrantly activated by mutation of genes encoding EGFR, KRAS, and PIK3CA proteins. We sought to understand whether class IA PI3K enzymes represent rational therapeutic targets in cells of non-squamous lung cancers by exploring pharmacological and genetic inhibitors of PI3K enzymes in a non-small cell lung cancer (NSCLC) cell line system. We found that class IA PI3K enzymes were expressed in all cell lines tested, but treatment of NSCLC lines with isoform-selective inhibitors (A66, TGX-221, CAL-101 and IC488743) had little effect on cell proliferation or prolonged inhibition of AKT activity. Inhibitory pharmacokinetic and pharmacodynamic responses were observed using these agents at non-isoform selective concentrations and with the pan-class I (ZSTK474) agent. Response to pharmacological inhibition suggested that PI3K isoforms may functionally compensate for one another thus limiting efficacy of single agent treatment. However, combination of ZSTK474 and an EGFR inhibitor (erlotinib) in NSCLC resistant to each single agent reduced cellular proliferation. These studies uncovered unanticipated cellular responses to PI3K isoform inhibition in NSCLC that does not correlate with PI3K mutations, suggesting that patients bearing tumors with wildtype EGFR and KRAS are unlikely to benefit from inhibitors of single isoforms but may respond to pan-isoform inhibition.

  4. Enterococcus faecalis infection activates phosphatidylinositol 3-kinase signaling to block apoptotic cell death in macrophages.

    PubMed

    Zou, Jun; Shankar, Nathan

    2014-12-01

    Apoptosis is an intrinsic immune defense mechanism in the host response to microbial infection. Not surprisingly, many pathogens have evolved various strategies to manipulate this important pathway to benefit their own survival and dissemination in the host during infection. To our knowledge, no attempts have been made to explore the host cell survival signals modulated by the bacterium Enterococcus faecalis. Here, we show for the first time that during early stages of infection, internalized enterococci can prevent host cell (RAW264.7 cells, primary macrophages, and mouse embryonic fibroblasts [MEFs]) apoptosis induced by a wide spectrum of proapoptotic stimuli. Activation of caspase 3 and cleavage of the caspase 3 substrate poly(ADP-ribose) polymerase were inhibited in E. faecalis-infected cells, indicating that E. faecalis protects macrophages from apoptosis by inhibiting caspase 3 activation. This antiapoptotic activity in E. faecalis-infected cells was dependent on the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, which resulted in the increased expression of the antiapoptotic factor Bcl-2 and decreased expression of the proapoptotic factor Bax. Further analysis revealed that active E. faecalis physiology was important for inhibition of host cell apoptosis, and this feature seemed to be a strain-independent trait among E. faecalis isolates. Employing a mouse peritonitis model, we also determined that cells collected from the peritoneal lavage fluid of E. faecalis-infected mice showed reduced levels of apoptosis compared to cells from uninfected mice. These results show early modulation of apoptosis during infection and have important implications for enterococcal pathogenesis.

  5. Phosphatidylinositol 3-kinase/Akt signaling pathway mediates acupuncture-induced dopaminergic neuron protection and motor function improvement in a mouse model of Parkinson's disease.

    PubMed

    Kim, Seung-Nam; Kim, Seung-Tae; Doo, Ah-Reum; Park, Ji-Yeun; Moon, Woongjoon; Chae, Younbyoung; Yin, Chang Shik; Lee, Hyejung; Park, Hi-Joon

    2011-10-01

    It has been reported that acupuncture treatment reduced dopaminergic neuron degeneration in Parkinson's disease (PD) models. However, the mechanistic pathways underlying, such neuroprotection, are poorly understood. Here, we investigated the effects and the underlying mechanism of acupuncture in a mouse model of PD using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). First, we observed that MPTP-induced impairment of Akt activation, but not MPTP-induced c-Jun activation, was effectively restored by acupuncture treatment in the substantia nigra. Furthermore, we demonstrated for the first time that the brain-specific blockade of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, by intranasal administration of LY294002, a specific inhibitor of PI3K/Akt signaling pathway, significantly blocked acupuncture-induced dopaminergic neuron protection and motor function improvement. Our results provide evidence that PI3K/Akt signaling pathway may play a central role in the mechanism underlying acupuncture-induced benefits in Parkinsonian mice.

  6. Echinacea purpurea root extract inhibits TNF release in response to Pam3Csk4 in a phosphatidylinositol-3-kinase dependent manner.

    PubMed

    Fast, David J; Balles, John A; Scholten, Jeffrey D; Mulder, Timothy; Rana, Jatinder

    2015-10-01

    Polysaccharides derived from Echinacea have historically been shown to be immunostimulatory. We describe in this work however the anti-inflammatory effect of a water extract of Echinacea purpurea roots (EPRW) that inhibited Pam3Csk4 stimulated production of TNFα by human monocytic THP-1 cells. The polyphenols and alkylamides typically found in Echinacea extracts were absent in EPRW suggesting that the anti-inflammatory component(s) was a polysaccharide. This anti-inflammatory activity was shown to be mediated by the phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway as chemical inhibition of PI3K abolished the EPRW anti-inflammatory effect. Demonstration of phosphorylation of Akt and ribosomal S6 proteins, downstream targets of PI3K confirmed EPRW-mediated activation of this pathway. In conclusion, this observation suggests that non-alkylamide/non-polyphenolic phytochemicals from Echinacea may contribute in part to some of the anti-inflammatory therapeutic effects such as reduced severity of symptoms that have been observed in vivo in the treatment of upper respiratory tract infections with Echinacea. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Adiponectin Induces Oncostatin M Expression in Osteoblasts through the PI3K/Akt Signaling Pathway

    PubMed Central

    Su, Chen-Ming; Lee, Wei-Lin; Hsu, Chin-Jung; Lu, Ting-Ting; Wang, Li-Hong; Xu, Guo-Hong; Tang, Chih-Hsin

    2015-01-01

    Rheumatoid arthritis (RA), a common autoimmune disorder, is associated with a chronic inflammatory response and unbalanced bone metabolism within the articular microenvironment. Adiponectin, an adipokine secreted by adipocytes, is involved in multiple functions, including lipid metabolism and pro-inflammatory activity. However, the mechanism of adiponectin performance within arthritic inflammation remains unclear. In this study, we observed the effect of adiponectin on the expression of oncostatin M (OSM), a pro-inflammatory cytokine, in human osteoblastic cells. Pretreatment of cells with inhibitors of phosphatidylinositol 3-kinase (PI3K), Akt, and nuclear factor (NF)-κB reduced the adiponectin-induced OSM expression in osteoblasts. Stimulation of the cells with adiponectin increased phosphorylation of PI3K, Akt, and p65. Adiponectin treatment of osteoblasts increased OSM-luciferase activity and p65 binding to NF-κB on the OSM promoter. Our results indicate that adiponectin increased OSM expression via the PI3K, Akt, and NF-κB signaling pathways in osteoblastic cells, suggesting that adiponectin is a novel target for arthritis treatment. PMID:26712749

  8. Involvement of PI3K/Akt/TOR pathway in stretch-induced hypertrophy of myotubes.

    PubMed

    Sasai, Nobuaki; Agata, Nobuhide; Inoue-Miyazu, Masumi; Kawakami, Keisuke; Kobayashi, Kunihiko; Sokabe, Masahiro; Hayakawa, Kimihide

    2010-01-01

    Skeletal muscle cells are hypertrophied by mechanical stresses, but the underlying molecular mechanisms are not fully understood. Two signaling pathways, phosphatidylinositol 3-kinase (PI3K)/Akt to target of rapamycin (TOR) and extracellular signal-regulated kinase kinase (MEK) to extracellular signal-regulated kinase (ERK), have been proposed to be involved in muscle hypertrophy. In this study we examined the involvement of these pathways in primary cultures of chick skeletal myotubes subjected to passive cyclic stretching for 72 hours, a time that was sufficient to induce significant hypertrophy in our preparations. Hypertrophy was largely suppressed by wortmannin or rapamycin, inhibitors of PI3K or mTOR, respectively. Furthermore, phosphorylation of Akt was enhanced by stretching and suppressed by wortmannin. The MEK inhibitor, U0126, exerted a minimal influence on stretch-induced hypertrophy. We found that cyclic stretching of myotubes activates the PI3K/Akt/TOR pathway, resulting in muscle hypertrophy. The MEK/ERK pathway may contribute negatively to spontaneous hypertrophy.

  9. Sequential application of a cytotoxic nanoparticle and a PI3K inhibitor enhances antitumor efficacy

    PubMed Central

    Pandey, Ambarish; Goldman, Aaron; Sarangi, Sasmit; Sengupta, Poulomi; Phipps, Colin; Kopparam, Jawahar; Oh, Michael; Basu, Sudipta; Kohandel, Mohammad; Sengupta, Shiladitya

    2013-01-01

    Nanomedicines that preferentially deploy cytotoxic agents to tumors, and molecular targeted therapeutics that inhibit specific aberrant oncogenic drivers are emerging as the new paradigm for the management of cancer. While combination therapies are a mainstay of cancer chemotherapy, few studies have addressed the combination of nanomedicines and molecular targeted therapeutics. Furthermore, limited knowledge exists on the impact of sequencing of such therapeutics and nanomedicines on the antitumor outcome. Here we engineered a supramolecular cis-platinum nanoparticle, which induced apoptosis in breast cancer cells but also elicited pro-survival signaling via an epidermal growth factor receptor-phosphatidylinositol 3 kinase (PI3K) pathway. A combination of mathematical modeling and in vitro and in vivo validation using a pharmacological inhibitor of PI3K, PI828, demonstrate that administration of PI828 following treatment with the supramolecular cis-platinum nanoparticle results in enhanced antitumor efficacy in breast cancer as compared with when the sequence is reversed or when the two treatments are administered simultaneously. This study addresses, for the first time, the impact of drug sequencing in the case of a combination of a nanomedicine and a targeted therapeutic. Furthermore, our results indicate that a rational combination of cis-platinum nanoparticles and a PI3K-targeted therapeutic can emerge as a potential therapy for breast cancer. PMID:24121494

  10. PI3K signaling in the pathogenesis of obesity: The cause and the cure.

    PubMed

    Beretta, Martina; Bauer, Michael; Hirsch, Emilio

    2015-05-01

    With the steady rise in the incidence of obesity and its associated comorbidities, in the last decades research aimed at understanding molecular mechanisms that control body weight has gained new interest. Fat gain is frequently associated with chronic adipose tissue inflammation and with peripheral as well as central metabolic derangements, resulting in an impaired hypothalamic regulation of energy homeostasis. Recent attention has focused on the role of phosphatidylinositol 3-kinase (PI3K) in both immune and metabolic response pathways, being involved in the pathophysiology of obesity and its associated metabolic diseases. In this review, we focus on distinct PI3K isoforms, especially class I PI3Ks, mediating inflammatory cells recruitment to the enlarged fat as well as intracellular responses to key hormonal regulators of fat storage, both in adipocytes and in the central nervous system. This integrated view of PI3K functions may ultimately help to develop new therapeutic interventions for the treatment of obesity. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Mutation of genes of the PI3K/AKT pathway in breast cancer supports their potential importance as biomarker for breast cancer aggressiveness.

    PubMed

    Tserga, Aggeliki; Chatziandreou, Ilenia; Michalopoulos, Nicolaos V; Patsouris, Efstratios; Saetta, Angelica A

    2016-07-01

    Deregulation of phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway is closely associated with cancer development and cancer progression. PIK3CA, AKT1, and PTEN are the fundamental molecules of the PI3K/AKT pathway with increased mutation rates in cancer cases leading to aberrant regulation of the pathway. Even though molecular alterations of the PI3K/AKT pathway have been studied in breast cancer, correlations between specific molecular alterations and clinicopathological features remain contradictory. In this study, we examined mutations of the PI3K/AKT pathway in 75 breast carcinomas using high-resolution melting analysis and pyrosequencing, in parallel with analysis of relative expression of PIK3CA and AKT2 genes. Mutations of PIK3CA were found in our cohort in 21 cases (28 %), 10 (13 %) in exon 9 and 11(15 %) in exon 20. Mutation frequency of AKT1 and PTEN genes was 4 and 3 %, respectively. Overall, alterations in the PI3K/AKT signaling cascade were detected in 35 % of the cases. Furthermore, comparison of 50 breast carcinomas with adjacent normal tissues showed elevated PIK3CA messenger RNA (mRNA) levels in 18 % of tumor cases and elevated AKT2 mRNA levels in 14 %. Our findings, along with those of previous studies, underline the importance of the PI3K/AKT pathway components as potential biomarkers for breast carcinogenesis.

  12. Quercetin regresses Dalton's lymphoma growth via suppression of PI3K/AKT signaling leading to upregulation of p53 and decrease in energy metabolism.

    PubMed

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2015-01-01

    Various oncogenes are associated with deregulation in cell proliferation, apoptosis, and cell survival, which ultimately cause cancerous growth. Phosphatidylinositol 3-kinase (PI3K) mediated signaling plays a key role in malignant transformation. Cell proliferation and cell survival of tumor cell are induced by hyper activation of PI3K, AKT1, glycolytic enzyme LDH-A, and inactivation of tumor suppressor gene p53. Dietary flavonoids such as quercetin are considered a powerful modulator of different cellular signaling pathways. The present study is focused on the role of quercetin on regulation of PI3K/AKT pathways in Dalton's lymphoma mice. Effect of quercetin was analyzed in ascite cells in terms of cell viability, glycolytic metabolism as well as expression, and level of PI3K (regulatory and catalytic subunit), AKT1, and p53 using standard methods. Results reflect hyperactivation of PI3K signaling in ascite cells of Dalton's lymphoma mice, leading to activation of AKT1 and inactivation of p53. Quercetin modulates the pathway toward suppression of lymphoma. Glycolytic metabolism was also downregulated by quercetin. Its tumor suppressor activity was confirmed by morphological parameters and longevity of mice. The findings suggest that quercetin may contribute to lymphoma prevention by downregulating PI3K-AKT1-p53 pathway as well as by glycolytic metabolism.

  13. Phosphatidylinositol 3-kinase activity and asymmetrical accumulation of F-actin are necessary for establishment of cell polarity in the early development of monospores from the marine red alga Porphyra yezoensis.

    PubMed

    Li, Lin; Saga, Naotsune; Mikami, Koji

    2008-01-01

    The polarized distribution of F-actin is important in providing the driving force for directional migration in mammalian leukocytes and Dictyostelium cells, in which compartmentation of phosphatidylinositol 3-kinase (PI3K) and phosphatidylinositol phosphatase is critical for the establishment of cell polarity. Since monospores from the red alga Porphyra yezoensis are a real example of migrating plant cells, the involvement of the cytoskeleton and PI3K was investigated during their early development. Our results indicate that the asymmetrical localization of F-actin at the leading edge is fixed by the establishment of the anterior-posterior axis in migrating monospores, which is PI3K-dependent and protein synthesis-independent. After migration, monospores adhere to the substratum and then become upright, developing into multicellular thalli via the establishment of the apical-basal axis. In this process, F-actin usually accumulates at the bottom of the basal cell and development after migration requires new protein synthesis. These findings suggest that the establishment of anterior-posterior and apical-basal axes are differentially regulated during the early development of monospores. Our results also indicate that PI3K-dependent F-actin asymmetry is evolutionally conserved in relation to the establishment of cell polarity in migrating eukaryotic cells.

  14. Architecture and dynamics of the autophagic phosphatidylinositol 3-kinase complex

    PubMed Central

    Baskaran, Sulochanadevi; Carlson, Lars-Anders; Stjepanovic, Goran; Young, Lindsey N; Kim, Do Jin; Grob, Patricia; Stanley, Robin E; Nogales, Eva; Hurley, James H

    2014-01-01

    The class III phosphatidylinositol 3-kinase complex I (PI3KC3-C1) that functions in early autophagy consists of the lipid kinase VPS34, the scaffolding protein VPS15, the tumor suppressor BECN1, and the autophagy-specific subunit ATG14. The structure of the ATG14-containing PI3KC3-C1 was determined by single-particle EM, revealing a V-shaped architecture. All of the ordered domains of VPS34, VPS15, and BECN1 were mapped by MBP tagging. The dynamics of the complex were defined using hydrogen–deuterium exchange, revealing a novel 20-residue ordered region C-terminal to the VPS34 C2 domain. VPS15 organizes the complex and serves as a bridge between VPS34 and the ATG14:BECN1 subcomplex. Dynamic transitions occur in which the lipid kinase domain is ejected from the complex and VPS15 pivots at the base of the V. The N-terminus of BECN1, the target for signaling inputs, resides near the pivot point. These observations provide a framework for understanding the allosteric regulation of lipid kinase activity. DOI: http://dx.doi.org/10.7554/eLife.05115.001 PMID:25490155

  15. Endosomal Phosphatidylinositol 3-Kinase Is Essential for Canonical GPCR Signaling.

    PubMed

    Uchida, Yasunori; Rutaganira, Florentine U; Jullié, Damien; Shokat, Kevan M; von Zastrow, Mark

    2017-01-01

    G protein-coupled receptors (GPCRs), the largest family of signaling receptors, are critically regulated by endosomal trafficking, suggesting that endosomes might provide new strategies for manipulating GPCR signaling. Here we test this hypothesis by focusing on class III phosphatidylinositol 3-kinase (Vps34), which is an essential regulator of endosomal trafficking. We verify that Vps34 is required for recycling of the β2-adrenoceptor (β2AR), a prototypical GPCR, and then investigate the effects of Vps34 inhibition on the canonical cAMP response elicited by β2AR activation. Vps34 inhibition impairs the ability of cells to recover this response after prolonged activation, which is in accord with the established role of recycling in GPCR resensitization. In addition, Vps34 inhibition also attenuates the short-term cAMP response, and its effect begins several minutes after initial agonist application. These results establish Vps34 as an essential determinant of both short-term and long-term canonical GPCR signaling, and support the potential utility of the endosomal system as a druggable target for signaling.

  16. P-REX1 creates a positive feedback loop to activate growth factor receptor, PI3K/AKT and MEK/ERK signaling in breast cancer.

    PubMed

    Dillon, L M; Bean, J R; Yang, W; Shee, K; Symonds, L K; Balko, J M; McDonald, W H; Liu, S; Gonzalez-Angulo, A M; Mills, G B; Arteaga, C L; Miller, T W

    2015-07-23

    Phosphatidylinositol 3-kinase (PI3K) promotes cancer cell survival, migration, growth and proliferation by generating phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the inner leaflet of the plasma membrane. PIP3 recruits pleckstrin homology domain-containing proteins to the membrane to activate oncogenic signaling cascades. Anticancer therapeutics targeting the PI3K/AKT/mTOR (mammalian target of rapamycin) pathway are in clinical development. In a mass spectrometric screen to identify PIP3-regulated proteins in breast cancer cells, levels of the Rac activator PIP3-dependent Rac exchange factor-1 (P-REX1) increased in response to PI3K inhibition, and decreased upon loss of the PI3K antagonist phosphatase and tensin homolog (PTEN). P-REX1 mRNA and protein levels were positively correlated with ER expression, and inversely correlated with PI3K pathway activation in breast tumors as assessed by gene expression and phosphoproteomic analyses. P-REX1 increased activation of Rac1, PI3K/AKT and MEK/ERK signaling in a PTEN-independent manner, and promoted cell and tumor viability. Loss of P-REX1 or inhibition of Rac suppressed PI3K/AKT and MEK/ERK, and decreased viability. P-REX1 also promoted insulin-like growth factor-1 receptor activation, suggesting that P-REX1 provides positive feedback to activators upstream of PI3K. In support of a model where PIP3-driven P-REX1 promotes both PI3K/AKT and MEK/ERK signaling, high levels of P-REX1 mRNA (but not phospho-AKT or a transcriptomic signature of PI3K activation) were predictive of sensitivity to PI3K inhibitors among breast cancer cell lines. P-REX1 expression was highest in estrogen receptor-positive breast tumors compared with many other cancer subtypes, suggesting that neutralizing the P-REX1/Rac axis may provide a novel therapeutic approach to selectively abrogate oncogenic signaling in breast cancer cells.

  17. Balanced Interactions between Lyn, the p85α Regulatory Subunit of Class IA Phosphatidylinositol-3-Kinase, and SHIP Are Essential for Mast Cell Growth and Maturation▿

    PubMed Central

    Ma, Peilin; Vemula, Sasidhar; Munugalavadla, Veerendra; Chen, Jinbiao; Sims, Emily; Borneo, Jovencio; Kondo, Takako; Ramdas, Baskar; Mali, Raghuveer Singh; Li, Shuo; Hashino, Eri; Takemoto, Clifford; Kapur, Reuben

    2011-01-01

    The growth and maturation of bone marrow-derived mast cells (BMMCs) from precursors are regulated by coordinated signals from multiple cytokine receptors, including KIT. While studies conducted using mutant forms of these receptors lacking the binding sites for Src family kinases (SFKs) and phosphatidylinositol-3-kinase (PI3K) suggest a role for these signaling molecules in regulating growth and survival, how complete loss of these molecules in early BMMC progenitors (MCps) impacts maturation and growth during all phases of mast cell development is not fully understood. We show that the Lyn SFK and the p85α subunit of class IA PI3K play opposing roles in regulating the growth and maturation of BMMCs in part by regulating the level of PI3K. Loss of Lyn in BMMCs results in elevated PI3K activity and hyperactivation of AKT, which accelerates the rate of BMMC maturation due in part to impaired binding and phosphorylation of SHIP via Lyn's unique domain. In the absence of Lyn's unique domain, BMMCs behave in a manner similar to that of Lyn- or SHIP-deficient BMMCs. Importantly, loss of p85α in Lyn-deficient BMMCs not only represses the hyperproliferation associated with the loss of Lyn but also represses their accelerated maturation. The accelerated maturation of BMMCs due to loss of Lyn is associated with increased expression of microphthalmia-associated transcription factor (Mitf), which is repressed in MCps deficient in the expression of both Lyn and p85α relative to controls. Our results demonstrate a crucial interplay of Lyn, SHIP, and p85α in regulating the normal growth and maturation of BMMCs, in part by regulating the activation of AKT and the expression of Mitf. PMID:21791602

  18. Activity of the novel dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor NVP-BEZ235 against T-cell acute lymphoblastic leukemia.

    PubMed

    Chiarini, Francesca; Grimaldi, Cecilia; Ricci, Francesca; Tazzari, Pier Luigi; Evangelisti, Camilla; Ognibene, Andrea; Battistelli, Michela; Falcieri, Elisabetta; Melchionda, Fraia; Pession, Andrea; Pagliaro, Pasqualepaolo; McCubrey, James A; Martelli, Alberto M

    2010-10-15

    Recent findings have highlighted that constitutively active phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling is a common feature of T-cell acute lymphoblastic leukemia (T-ALL), where it upregulates cell proliferation, survival, and drug resistance. These observations lend compelling weight to the application of PI3K/Akt/mTOR inhibitors in the therapy of T-ALL. Here, we have analyzed the therapeutic potential of the novel dual PI3K/mTOR inhibitor NVP-BEZ235, an orally bioavailable imidazoquinoline derivative, which has entered clinical trials for solid tumors, on both T-ALL cell lines and patient samples. NVP-BEZ235 was cytotoxic to a panel of T-ALL cell lines as determined by MTT assays. NVP-BEZ235 treatment resulted in cell cycle arrest and apoptosis. Western blots showed a dose- and time-dependent dephosphorylation of Akt and mTORC1 downstream targets in response to NVP-BEZ235. Remarkably, NVP-BEZ235 targeted the side population of both T-ALL cell lines and patient lymphoblasts, which might correspond to leukemia-initiating cells, and synergized with chemotherapeutic agents (cyclophosphamide, cytarabine, dexamethasone) currently used for treating T-ALL patients. NVP-BEZ235 reduced chemoresistance to vincristine induced in Jurkat cells by coculturing with MS-5 stromal cells, which mimic the bone marrow microenvironment. NVP-BEZ235 was cytotoxic to T-ALL patient lymphoblasts displaying pathway activation, where the drug dephosphorylated eukaryotic initiation factor 4E-binding protein 1, at variance with rapamycin. Taken together, our findings indicate that longitudinal inhibition at two nodes of the PI3K/Akt/mTOR network with NVP-BEZ235, either alone or in combination with chemotherapeutic drugs, may be an efficient treatment of those T-ALLs that have aberrant upregulation of this signaling pathway for their proliferation and survival. ©2010 AACR.

  19. Neurotoxicity of developmental hypothyroxinemia and hypothyroidism in rats: Impairments of long-term potentiation are mediated by phosphatidylinositol 3-kinase signaling pathway

    SciTech Connect

    Wang, Yi; Wei, Wei; Wang, Yuan; Dong, Jing; Song, Binbin; Min, Hui; Teng, Weiping; Chen, Jie

    2013-09-01

    Neurotoxicity of iodine deficiency-induced hypothyroidism during developmental period results in serious impairments of brain function, such as learning and memory. These impairments are largely irreversible, and the underlying mechanisms remain unclear. In addition to hypothyroidism, iodine deficiency may cause hypothyroxinemia, a relatively subtle form of thyroid hormone deficiency. Neurotoxicity of developmental hypothyroxinemia also potentially impairs learning and memory. However, more direct evidence of the associations between developmental hypothyroxinemia and impairments of learning and memory should be provided, and the underlying mechanisms remain to be elucidated. Thus, in the present study, we investigated the effects of developmental hypothyroxinemia and hypothyroidism on long-term potentiation (LTP), a widely accepted cellular model of learning and memory, in the hippocampal CA1 region. The activation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway – a pathway closely associated with synaptic plasticity and learning and memory – was also investigated. Wistar rats were treated with iodine deficient diet or methimazole (MMZ) to induce developmental hypothyroxinemia or hypothyroidism. The results showed that developmental hypothyroxinemia caused by mild iodine deficiency and developmental hypothyroidism caused by severe iodine deficiency or MMZ significantly reduced the field-excitatory postsynaptic potential (f-EPSP) slope and the population spike (PS) amplitude. Decreased activation of the PI3K signaling pathway was also observed in rats subjected to developmental hypothyroxinemia or hypothyroidism. Our results may support the hypothesis that neurotoxicity of both developmental hypothyroxinemia and hypothyroidism causes damages to learning and memory. Our results also suggest that decreased activation of the PI3K signaling pathway may contribute to impairments of LTP caused by neurotoxicity of both developmental hypothyroxinemia and

  20. Dexmedetomidine increases the activity of excitatory amino acid transporter type 3 expressed in Xenopus oocytes: the involvement of protein kinase C and phosphatidylinositol 3-kinase.

    PubMed

    Do, Sang-Hwan; Park, Seong-Joo; Shin, Hyun-Jung; Paik, Hye-Sun; Zuo, Zhiyi; Yoon, Hea-Jo; Ryu, Jung-Hee

    2014-09-05

    Dexmedetomidine, an α2 adrenergic agonist, has neuroprotective and anticonvulsant properties in addition to its sedative and anxiolytic effects. We hypothesized that dexmedetomidine would increase the activity of excitatory amino acid transporter type 3 (EAAT3) and that this effect would involve protein kinase C (PKC) and phosphatidylinositol 3-kinase (PI3K), two protein kinases known to regulate EAAT3 activity. EAAT3 was expressed in Xenopus oocytes by injecting its mRNA. Two-electrode voltage clamping was used to record membrane currents before, during, and after application of 30 μM l-glutamate in the presence of 0.1-30 nM dexmedetomidine. Dexmedetomidine-treated oocytes were also exposed to a PKC activator (phorbol-12-myristate-13-acetate [PMA]), PKC inhibitors (chelerythrine, staurosporine, and calphostin C), and PI3K inhibitors (wortmannin and LY294002) before current measurement. Dexmedetomidine application resulted in a concentration-dependent increase in the EAAT3 activity in response to l-glutamate. The kinetic study showed that dexmedetomidine significantly increased the Vmax without changing Km. Treatment of oocytes with PMA significantly increased transporter currents compared with controls, but treatment with dexmedetomidine plus PMA did not further increase the response compared with PMA or dexmedetomidine alone. In addition, pre-treatment of oocytes with PKC inhibitors and PI3K inhibitors significantly abolished the dexmedetomidine-enhanced EAAT3 activity. These results suggest that dexmedetomidine increases the activity of EAAT3 expressed in Xenopus oocytes. PKC and PI3K seem to mediate this effect. These findings may explain the neuroprotective and anticonvulsant effects of dexmedetomidine.

  1. Recombinant Human Erythropoietin Augments Neovascularization Responses in a Neonatal Rat Model of Premature Brain Damage by Phosphatidylinositol 3 Kinase/Akt Pathway.

    PubMed

    Yu, Da-Fan; Zhu, Li-Hua; Jiang, Li

    2017-04-05

    Recombinant human-erythropoietin (rh-EPO) has therapeutic efficacy for premature infants with brain damage during the active rehabilitation and anti-inflammation. In the present study, we found that the rh-EPO was related to the promotion of neovascularization. Our aim was to investigate whether rh-EPO augments neovascularization in the neonatal rat model of premature brain damage through the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) signaling pathway. Postnatal day 5 (PD5), rats underwent permanent ligation of the right common carotid artery and were exposed to hypoxia for 2 h. All the rat pups were randomized into five groups as follows: (1) control group; (2) hypoxia-ischemic (HI) group; (3) HI + LY294002 group; (4) HI + rh-EPO group; and (5) HI + rh-EPO + LY294002 group. The phospho-Akt protein was tested 90 min after the whole operation, and CD34, vascular endothelial growth factor receptor 2 (VEGFR2), and vascular endothelial growth factor (VEGF) were also tested 2 days after the whole operation. In the hypoxic and ischemic zone of the premature rat brain, the rh-EPO induced CD34+ cells to immigrate to the HI brain zone (P < 0.05) and also upregulated the VEGFR2 protein expression (P < 0.05) and VEGF mRNA level (P < 0.05) through the PI3K/Akt (P < 0.05) signaling pathway when compared with other groups. The rh-EPO treatment augments neovascularization responses in the neonatal rat model of premature brain damage through the PI3K/Akt signaling pathway. Besides, the endogenous EPO may exist in the HI zone of rat brain and also has neovascularization function through the PI3K/Akt signaling pathway.

  2. Phosphatidylinositol 3-kinase p85 alpha regulatory subunit gene PIK3R1 haplotype is associated with body fat and serum leptin in a female twin population

    PubMed Central

    Jamshidi, Y.; Snieder, H.; Wang, X.; Pavitt, M. J.; Spector, T. D.; Carter, N. D.; O’Dell, S. D.

    2006-01-01

    Aims/hypothesis Phosphatidylinositol 3-kinase (PI3K) couples the leptin and insulin signalling pathways via IRS-1 and IRS-2. Hence, defective activation of PI3K could be a novel mechanism of peripheral leptin or insulin resistance. We investigated association of tagging SNPs (tSNPs) in the PI3K p85α regulatory subunit gene PIK3R1 with anthropometry, leptin, body fat and insulin sensitivity in a female twin population of European extraction. Methods Eight tSNPs were genotyped in 2778 women (mean age 47.4±12.5 years) from the St Thomas’ UK Adult Twin Registry (Twins UK). Results SNP rs1550805 was associated with serum leptin (P=0.028), BMI (P=0.025), weight (P=0.019), total fat (P=0.004), % total fat (P=0.002), waist (P=0.025), central fat (P=0.005) and % central fat (P=0.005). SNPs rs7713645 and rs7709243 were associated with BMI (P=0.020; P=0.029), rs7709243 with weight, total and central fat, (P=0.026; P=0.031; P=0.023) and both SNPs with fasting glucose (P=0.003; P=0.001) and glucose 2h post OGTT (P=0.023; P=0.007). Haplotype 222 (freq. 7.2%) showed higher serum leptin (P=0.007) and body fat measures (Ps≤0.001) and haplotype 221 (freq. 38.7%) showed higher fasting and 2h-glucose (P=0.035; P=0.021), compared with the most common haplotype 111 (freq. 45.5%). Conclusions/interpretation Association of the PIK3R1 SNP rs1550805 with serum leptin and body fat may reflect diminished ability of PI3K to signal via IRS-1 or IRS-2 in response to leptin. PMID:17016694

  3. Accumulation of Splice Variants and Transcripts in Response to PI3K Inhibition in T Cells

    PubMed Central

    Riedel, Alice; Mofolo, Boitumelo; Avota, Elita; Schneider-Schaulies, Sibylle; Meintjes, Ayton; Mulder, Nicola; Kneitz, Susanne

    2013-01-01

    Background Measles virus (MV) causes T cell suppression by interference with phosphatidylinositol-3-kinase (PI3K) activation. We previously found that this interference affected the activity of splice regulatory proteins and a T cell inhibitory protein isoform was produced from an alternatively spliced pre-mRNA. Hypothesis Differentially regulated and alternatively splice variant transcripts accumulating in response to PI3K abrogation in T cells potentially encode proteins involved in T cell silencing. Methods To test this hypothesis at the cellular level, we performed a Human Exon 1.0 ST Array on RNAs isolated from T cells stimulated only or stimulated after PI3K inhibition. We developed a simple algorithm based on a splicing index to detect genes that undergo alternative splicing (AS) or are differentially regulated (RG) upon T cell suppression. Results Applying our algorithm to the data, 9% of the genes were assigned as AS, while only 3% were attributed to RG. Though there are overlaps, AS and RG genes differed with regard to functional regulation, and were found to be enriched in different functional groups. AS genes targeted extracellular matrix (ECM)-receptor interaction and focal adhesion pathways, while RG genes were mainly enriched in cytokine-receptor interaction and Jak-STAT. When combined, AS/RG dependent alterations targeted pathways essential for T cell receptor signaling, cytoskeletal dynamics and cell cycle entry. Conclusions PI3K abrogation interferes with key T cell activation processes through both differential expression and alternative splicing, which together actively contribute to T cell suppression. PMID:23383294

  4. Arecoline-induced phosphorylated p53 and p21(WAF1) protein expression is dependent on ATM/ATR and phosphatidylinositol-3-kinase in clone-9 cells.

    PubMed

    Chou, Wen-Wen; Guh, Jinn-Yuh; Tsai, Jung-Fa; Hwang, Chi-Ching; Chiou, Shean-Jaw; Chuang, Lea-Yea

    2009-06-01

    Betel-quid use is associated with liver cancer whereas its constituent arecoline is cytotoxic, genotoxic, and induces p53-dependent p21(WAF1) protein expression in Clone-9 cells (rat hepatocytes). The ataxia telangiectasia mutated (ATM)/rad3-related (ATR)-p53-p21(WAF1) and the phosphatidylinositol-3-kinase (PI3K)-mammalian target of rapamycin (mTOR) pathways are involved in the DNA damage response and the pathogenesis of cancers. Thus, we studied the role of ATM/ATR and PI3K in arecoline-induced p53 and p21(WAF1) protein expression in Clone-9 cells. We found that arecoline (0.5 mM) activated the ATM/ATR kinase at 30 min. The arecoline-activated ATM/ATR substrate contained p-p53Ser15. Moreover, arecoline only increased the levels of the p-p53Ser6, p-p53Ser15, and p-p53Ser392 phosphorylated p53 isoforms among the known isoforms. ATM shRNA attenuated arecoline-induced p-p53Ser15 and p21(WAF1) at 24 h. Arecoline (0.5 mM) increased phosphorylation levels of p-AktSer473 and p-mTORSer2448 at 30-60 min. Dominant-negative PI3K plasmids attenuated arecoline-induced p21(WAF1), but not p-p53Ser15, at 24 h. Rapamycin attenuated arecoline-induced phosphrylated p-p53Ser15, but not p21(WAF1), at 24 h. ATM shRNA, but not dominant-negative PI3K plasmids, attenuated arecoline-induced p21(WAF1) gene transcription. We conclude that arecoline activates the ATM/ATR-p53-p21(WAF1) and the PI3K/Akt-mTOR-p53 pathways in Clone-9 cells. Arecoline-induced phosphorylated p-p53Ser15 expression is dependent on ATM whereas arecoline-induced p21(WAF1) protein expression is dependent on ATM and PI3K. Moreover, p21(WAF1) gene is transcriptionally induced by arecoline-activated ATM. (c) 2009 Wiley-Liss, Inc.

  5. Managing postmenopausal women with hormone receptor-positive advanced breast cancer who progress on endocrine therapies with inhibitors of the PI3K pathway.

    PubMed

    Brufsky, Adam M

    2014-01-01

    Although endocrine therapies that interfere with estrogen receptor (ER)-mediated signaling have revolutionized the management of postmenopausal women with hormone receptor-positive (HR+) breast cancer (BC), long-term management of these patients is suboptimal because of the eventual emergence of endocrine resistance. Intense research has elucidated a number of targets that act downstream or upstream of the ER, as well as those that crosstalk with the ER; however, clinical validation of inhibiting specific targets to overcome endocrine resistance has been lacking. The phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway has been implicated to mediate endocrine resistance, and a number of novel agents that target this pathway are in early- and late-stage clinical trials. Recently, everolimus, an inhibitor of mTOR, a critical component of the PI3K/AKT/mTOR pathway, in combination with endocrine therapy, was shown to prolong progression-free survival with a manageable adverse-event profile in postmenopausal patients with HR+ BC. Bolstered by the safety and efficacy observed with concomitant inhibition of the ER and the PI3K/mTOR pathway and the validation of dual inhibition approach in managing postmenopausal patients with HR+ BC, a number of novel agents that inhibit PI3K (pan-PI3K inhibitors) or PI3K and mTOR (dual PI3K/mTOR) are being evaluated in clinical trials. Thus, mTOR inhibitors have provided the much-needed ammunition to oncologists who manage postmenopausal women with BC and have paved the way for the development of novel therapies that target the PI3K/mTOR pathway. Use of these novel therapies in managing postmenopausal women with BC, in combination with endocrine therapies, is expected to improve overall outcomes by overcoming endocrine resistance. © 2014 Wiley Periodicals, Inc.

  6. The p110δ subunit of PI3K regulates bone marrow-derived eosinophil trafficking and airway eosinophilia in allergen-challenged mice.

    PubMed

    Kang, Bit Na; Ha, Sung Gil; Ge, Xiao Na; Reza Hosseinkhani, M; Bahaie, Nooshin S; Greenberg, Yana; Blumenthal, Malcolm N; Puri, Kamal D; Rao, Savita P; Sriramarao, P

    2012-06-01

    Trafficking and recruitment of eosinophils during allergic airway inflammation is mediated by the phosphatidylinositol 3-kinase (PI3K) family of signaling molecules. The role played by the p110δ subunit of PI3K (PI3K p110δ) in regulating eosinophil trafficking and recruitment was investigated using a selective pharmacological inhibitor (IC87114). Treatment with the PI3K p110δ inhibitor significantly reduced murine bone marrow-derived eosinophil (BM-Eos) adhesion to VCAM-1 as well as ICAM-1 and inhibited activation-induced changes in cell morphology associated with reduced Mac-1 expression and aberrant cell surface localization/distribution of Mac-1 and α4. Infused BM-Eos demonstrated significantly decreased rolling and adhesion in inflamed cremaster muscle microvessels of mice treated with IC87114 compared with vehicle-treated mice. Furthermore, inhibition of PI3K p110δ significantly attenuated eotaxin-1-induced BM-Eos migration and prevented eotaxin-1-induced changes in the cytoskeleton and cell morphology. Knockdown of PI3K p110δ with siRNA in BM-Eos resulted in reduced rolling, adhesion, and migration, as well as inhibition of activation-induced changes in cell morphology, validating its role in regulating trafficking and migration. Finally, in a mouse model of cockroach antigen-induced allergic airway inflammation, oral administration of the PI3K p110δ inhibitor significantly inhibited airway eosinophil recruitment, resulting in attenuation of airway hyperresponsiveness in response to methacholine, reduced mucus secretion, and expression of proinflammatory molecules (found in inflammatory zone-1 and intelectin-1). Overall, these findings indicate the important role played by PI3K p110δ in mediating BM-Eos trafficking and migration by regulating adhesion molecule expression and localization/distribution as well as promoting changes in cell morphology that favor recruitment during inflammation.

  7. The p110δ subunit of PI3K regulates bone marrow-derived eosinophil trafficking and airway eosinophilia in allergen-challenged mice

    PubMed Central

    Kang, Bit Na; Ha, Sung Gil; Ge, Xiao Na; Reza Hosseinkhani, M.; Bahaie, Nooshin S.; Greenberg, Yana; Blumenthal, Malcolm N.; Puri, Kamal D.; Rao, Savita P.

    2012-01-01

    Trafficking and recruitment of eosinophils during allergic airway inflammation is mediated by the phosphatidylinositol 3-kinase (PI3K) family of signaling molecules. The role played by the p110δ subunit of PI3K (PI3K p110δ) in regulating eosinophil trafficking and recruitment was investigated using a selective pharmacological inhibitor (IC87114). Treatment with the PI3K p110δ inhibitor significantly reduced murine bone marrow-derived eosinophil (BM-Eos) adhesion to VCAM-1 as well as ICAM-1 and inhibited activation-induced changes in cell morphology associated with reduced Mac-1 expression and aberrant cell surface localization/distribution of Mac-1 and α4. Infused BM-Eos demonstrated significantly decreased rolling and adhesion in inflamed cremaster muscle microvessels of mice treated with IC87114 compared with vehicle-treated mice. Furthermore, inhibition of PI3K p110δ significantly attenuated eotaxin-1-induced BM-Eos migration and prevented eotaxin-1-induced changes in the cytoskeleton and cell morphology. Knockdown of PI3K p110δ with siRNA in BM-Eos resulted in reduced rolling, adhesion, and migration, as well as inhibition of activation-induced changes in cell morphology, validating its role in regulating trafficking and migration. Finally, in a mouse model of cockroach antigen-induced allergic airway inflammation, oral administration of the PI3K p110δ inhibitor significantly inhibited airway eosinophil recruitment, resulting in attenuation of airway hyperresponsiveness in response to methacholine, reduced mucus secretion, and expression of proinflammatory molecules (found in inflammatory zone-1 and intelectin-1). Overall, these findings indicate the important role played by PI3K p110δ in mediating BM-Eos trafficking and migration by regulating adhesion molecule expression and localization/distribution as well as promoting changes in cell morphology that favor recruitment during inflammation. PMID:22427531

  8. Cold stress-induced brain injury regulates TRPV1 channels and the PI3K/AKT signaling pathway.

    PubMed

    Liu, Ying; Liu, Yunen; Jin, Hongxu; Cong, Peifang; Zhang, Yubiao; Tong, Changci; Shi, Xiuyun; Liu, Xuelei; Tong, Zhou; Shi, Lin; Hou, Mingxiao

    2017-09-01

    Transient receptor potential vanilloid 1 (TRPV1) is a nonselective cation channel that interacts with several intracellular proteins in vivo, including calmodulin and Phosphatidylinositol-3-Kinase/Protein Kinase B (PI3K/Akt). TRPV1 activation has been reported to exert neuroprotective effects. The aim of this study was to examine the impact of cold stress on the mouse brain and the underlying mechanisms of TRPV1 involvement. Adult male C57BL/6 mice were subjected to cold stress (4°C for 8h per day for 2weeks). The behavioral deficits of the mice were then measured using the Morris water maze. Expression levels of brain injury-related proteins and mRNA were measured by western blot, immunofluorescence or RT-PCR analysis. The mice displayed behavioral deficits, inflammation and changes in brain injury markers following cold stress. As expected, upregulated TRPV1 expression levels and changes in PI3K/Akt expression were found. The TRPV1 inhibitor reduced the levels of brain injury-related proteins and inflammation. These data suggest that cold stress can induce brain injury, possibly through TRPV1 activation and the PI3K/Akt signaling pathway. Suppression of inflammation by inhibition of TRPV1 and the PI3K/Akt pathway may be helpful to prevent cold stress-induced brain injury. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Aged black garlic extract inhibits HT29 colon cancer cell growth via the PI3K/Akt signaling pathway.

    PubMed

    Dong, Menghua; Yang, Guiqing; Liu, Hanchen; Liu, Xiaoxu; Lin, Sixiang; Sun, Dongning; Wang, Yishan

    2014-03-01

    Accumulating evidence indicates that aged black garlic extract (ABGE) may prove beneficial in preventing or inhibiting oncogenesis; however, the underlying mechanisms have not been fully elucidated. The present study aimed to investigate the effects of ABGE on the proliferation and apoptosis of HT29 colon cancer cells. Our results demonstrated that ABGE inhibited HT29 cell growth via the induction of apoptosis and cell cycle arrest. We further investigated the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signal transduction pathway and the molecular mechanisms underlying the ABGE-induced inhibition of HT29 cell proliferation. We observed that ABGE may regulate the function of the PI3K/Akt pathway through upregulating PTEN and downregulating Akt and p-Akt expression, as well as suppressing its downstream target, 70-kDa ribosomal protein S6 kinase 1, at the mRNA and protein levels. In conclusion, these findings suggest that the PI3K/Akt signal transduction pathway is crucial for the development of colon cancer. ABGE inhibited the growth and induced apoptosis in HT29 cells through the inhibition of the PI3K/Akt pathway, suggesting that ABGE may be effective in the prevention and treatment of colon cancer in humans.

  10. K-Ras promotes growth transformation and invasion of immortalized human pancreatic cells by Raf and phosphatidylinositol 3-kinase signaling.

    PubMed

    Campbell, Paul M; Groehler, Angela L; Lee, Kwang M; Ouellette, Michel M; Khazak, Vladimir; Der, Channing J

    2007-03-01

    Mutational activation of the K-Ras oncogene is well established as a key genetic step in the development and growth of pancreatic adenocarcinomas. However, the mechanism by which aberrant Ras signaling promotes uncontrolled pancreatic tumor cell growth remains to be fully elucidated. The recent use of primary human cells to study Ras-mediated oncogenesis provides important model cell systems to dissect this mechanism. We have used a model of telomerase-immortalized human pancreatic duct-derived cells (E6/E7/st) to study mechanisms of Ras growth transformation. First, we found that human papillomavirus E6 and E7 oncogenes, which block the function of the p53 and Rb tumor suppressors, respectively, and SV40 small t antigen were required to allow mutant K-Ras(12D) growth transformation. Second, K-Ras(12D) caused growth transformation in vitro, including enhanced growth rate and loss of density dependency for growth, anchorage independence, and invasion through reconstituted basement membrane proteins, and tumorigenic transformation in vivo. Third, we determined that the Raf, phosphatidylinositol 3-kinase (PI3K), and Ral guanine nucleotide exchange factor effector pathways were activated, although extracellular signal-regulated kinase (ERK) activity was not up-regulated persistently. Finally, pharmacologic inhibition of Raf/mitogen-activated protein kinase/ERK and PI3K signaling impaired K-Ras-induced anchorage-independent growth and invasion. In summary, our studies established, characterized, and validated E6/E7/st cells for the study of Ras-induced oncogenesis.

  11. Peroxynitrite activates glucose uptake in 3T3-L1 adipocytes through a PI3-K-dependent mechanism.

    PubMed

    Guzman-Grenfell, Alberto M; Garcia-Macedo, Rebeca; Gonzalez-Martinez, Marco T; Hicks, Juan Jose; Medina-Navarro, Rafael

    2005-01-01

    Peroxynitrite, the product of the reaction between *NO and O2*-, is a strong oxidant and nitrating molecule, and it has been recently consideredas a component of some important signaling pathways. Herein, we report the effect of peroxynitrite on glucose uptake in 3T3-L1 adipocytes. Peroxynitrite stimulated glucose uptake and this effect was inhibited by citochalasin B, indicating the participation of facilitated GLUT transporters. Peroxynitrite-induced glucose uptake was not related to intracellular ATP, nor to external or internal calcium, but it was inhibited by the phosphatidylinositol 3-kinase (PI3-K) inhibitor, wortmannin. Additionally, we also found that peroxynitrite did not activate the insulin receptor nor the PI3-K downstream signaling protein kinase B (PKB/Akt). The dose-dependent inhibitory action of wortmannin suggests that peroxynitrite activates glucose transport without affecting GLUT transporters translocation.

  12. PI3K Phosphorylation Is Linked to Improved Electrical Excitability in an In Vitro Engineered Heart Tissue Disease Model System

    PubMed Central

    Kana, Kujaany; Song, Hannah; Laschinger, Carol

    2015-01-01

    Myocardial infarction, a prevalent cardiovascular disease, is associated with cardiomyocyte cell death, and eventually heart failure. Cardiac tissue engineering has provided hopes for alternative treatment options, and high-fidelity tissue models for drug discovery. The signal transduction mechanisms relayed in response to mechanoelectrical (physical) stimulation or biochemical stimulation (hormones, cytokines, or drugs) in engineered heart tissues (EHTs) are poorly understood. In this study, an EHT model was used to elucidate the signaling mechanisms involved when insulin was applied in the presence of electrical stimulation, a stimulus that mimics functional heart tissue environment in vitro. EHTs were insulin treated, electrically stimulated, or applied in combination (insulin and electrical stimulation). Electrical excitability parameters (excitation threshold and maximum capture rate) were measured. Protein kinase B (AKT) and phosphatidylinositol-3-kinase (PI3K) phosphorylation revealed that insulin and electrical stimulation relayed electrical excitability through two separate signaling cascades, while there was a negative crosstalk between sustained activation of AKT and PI3K. PMID:26120935

  13. PI3K Phosphorylation Is Linked to Improved Electrical Excitability in an In Vitro Engineered Heart Tissue Disease Model System.

    PubMed

    Kana, Kujaany; Song, Hannah; Laschinger, Carol; Zandstra, Peter W; Radisic, Milica

    2015-09-01

    Myocardial infarction, a prevalent cardiovascular disease, is associated with cardiomyocyte cell death, and eventually heart failure. Cardiac tissue engineering has provided hopes for alternative treatment options, and high-fidelity tissue models for drug discovery. The signal transduction mechanisms relayed in response to mechanoelectrical (physical) stimulation or biochemical stimulation (hormones, cytokines, or drugs) in engineered heart tissues (EHTs) are poorly understood. In this study, an EHT model was used to elucidate the signaling mechanisms involved when insulin was applied in the presence of electrical stimulation, a stimulus that mimics functional heart tissue environment in vitro. EHTs were insulin treated, electrically stimulated, or applied in combination (insulin and electrical stimulation). Electrical excitability parameters (excitation threshold and maximum capture rate) were measured. Protein kinase B (AKT) and phosphatidylinositol-3-kinase (PI3K) phosphorylation revealed that insulin and electrical stimulation relayed electrical excitability through two separate signaling cascades, while there was a negative crosstalk between sustained activation of AKT and PI3K.

  14. BRAIN OVERGROWTH IN DISORDERS OF RTK-PI3K-AKT SIGNALING: A MOSAIC OF MALFORMATIONS

    PubMed Central

    Hevner, Robert F.

    2014-01-01

    Disorders of brain overgrowth are significant causes of intractable epilepsy, intellectual disability, autism, and other complex neurological problems. The pathology of these disorders is sometimes striking and characteristic, as in hemimegalencephaly, but can also be subtle, as in autism. Recent genetic studies have shown that many diverse forms of brain overgrowth are caused by de novo mutations that increase activity in the receptor tyrosine kinase (RTK)–phosphatidylinositol-3-kinase (PI3K)–AKT signaling pathway, a key mediator of signaling by growth factors in the developing brain, such as fibroblast growth factors. In cases where mutations arise in postzygotic embryos, brain regions exhibit mosaic pathology that reflects the distribution of mutant cells, ranging from focal cortical dysplasia to lobar or hemispheric overgrowth. In turn, the histopathology of these disorders is also remarkably varied. The common underlying mechanisms of RTK-PI3K-AKT overactivation suggest new possibilities for drugs that inhibit this pathway. PMID:25432429

  15. Imidazo[1,2-a]pyrazines as novel PI3K inhibitors.

    PubMed

    Martínez González, Sonia; Hernández, Ana Isabel; Varela, Carmen; Rodríguez-Arístegui, Sonsoles; Alvarez, Rosa María; García, Ana Belén; Lorenzo, Milagros; Rivero, Virginia; Oyarzabal, Julen; Rabal, Obdulia; Bischoff, James R; Albarrán, Maribel; Cebriá, Antonio; Alfonso, Patricia; Link, Wolfgang; Fominaya, Jesús; Pastor, Joaquín

    2012-03-01

    Phosphoinositide-3-kinase (PI3K) is an important target for cancer therapeutics due to the deregulation of its signaling pathway in a wide variety of human cancers. We describe herein a novel series of imidazo[1,2-a]pyrazines as PI3K inhibitors. Copyright © 2012 Elsevier Ltd. All rights reserved.

  16. Angiotensin II-induced delayed stimulation of phospholipase C gamma1 requires activation of both phosphatidylinositol 3-kinase gamma and tyrosine kinase in vascular myocytes.

    PubMed

    Rakotoarisoa, Lala; Carricaburu, Valérie; Leblanc, Catherine; Mironneau, Chantal; Mironneau, Jean; Macrez, Nathalie

    2006-01-01

    In vascular smooth muscles, angiotensin II (AII) has been reported to activate phospholipase C (PLC) and phosphatidylinositol 3-kinase (PI3K). We investigated the time-dependent effects of AII on both phosphatidylinositol 3,4,5-trisphosphate (PtdInsP3) and inositol phosphates (InsPs) accumulation in permeabilized microsomes from rat portal vein smooth muscle in comparison with those of noradrenaline (NA). AII stimulated an early production of PtdInsP3 (within 30 s) followed by a delayed production of InsPs (within 3-5 min), in contrast to NA which activated only a fast production of InsPs. The use of pharmacological inhibitors and antibodies raised against the PI3K and PLC isoforms expressed in portal vein smooth muscle showed that AII specifically activated PI3Kgamma and that this isoform was involved in the AII-induced stimulation of InsPs accumulation. NA-induced InsPs accumulation depended on PLCbeta1 activation whereas AII-induced InsPs accumulation depended on PLCgamma1 activation. AII-induced PLCgamma1 activation required both tyrosine kinase and PI3Kgamma since genistein and tyrphostin B48 (inhibitors of tyrosine kinase), LY294002 and wortmannin (inhibitors of PI3K) and anti-PI3Kgamma antibody abolished AII-induced stimulation of InsPs accumulation. Increased tyrosine phosphorylation of PLCgamma1 was only detected for long-lasting applications of AII and was suppressed by genistein. These data indicate that activation of both PI3Kgamma and tyrosine kinase is a prerequisite for AII-induced stimulation of PLCgamma1 in vascular smooth muscle and suggest that the sequential activation of the three enzymes may be responsible for the slow and long-lasting contraction induced by AII.

  17. The phosphoinositide 3-kinase pathway.

    PubMed

    Cantley, Lewis C

    2002-05-31

    Phosphorylated lipids are produced at cellular membranes during signaling events and contribute to the recruitment and activation of various signaling components. The role of phosphoinositide 3-kinase (PI3K), which catalyzes the production of phosphatidylinositol-3,4,5-trisphosphate, in cell survival pathways; the regulation of gene expression and cell metabolism; and cytoskeletal rearrangements are highlighted. The PI3K pathway is implicated in human diseases including diabetes and cancer, and understanding the intricacies of this pathway may provide new avenues for therapuetic intervention.

  18. Inhibition of net HepG2 cell apolipoprotein B secretion by the citrus flavonoid naringenin involves activation of phosphatidylinositol 3-kinase, independent of insulin receptor substrate-1 phosphorylation.

    PubMed

    Borradaile, Nica M; de Dreu, Linda E; Huff, Murray W

    2003-10-01

    The flavonoid naringenin improves hyperlipidemia and hyperglycemia in streptozotocin-treated rats. In HepG2 human hepatoma cells, naringenin inhibits apolipoprotein B (apoB) secretion primarily by inhibiting microsomal triglyceride transfer protein and enhances LDL receptor (LDLr)-mediated apoB-containing lipoprotein uptake. Phosphatidylinositol 3-kinase (PI3K) activation by insulin increases sterol regulatory element-binding protein (SREBP)-1 and LDLr expression and inhibits apoB secretion in hepatocytes. Thus, we determined whether naringenin activates this pathway. Insulin and naringenin induced PI3K-dependent increases in cytosolic and nuclear SREBP-1 and LDLr expression. Similar PI3K-mediated increases in SREBP-1 were observed in McA-RH7777 rat hepatoma cells, which express predominantly SREBP-1c. Reductions in HepG2 cell media apoB with naringenin were partially attenuated by wortmannin, whereas the effect of insulin was completely blocked. Both treatments reduced apoB100 secretion in wild-type and LDLr(-/-) mouse hepatocytes to the same extent. Insulin and naringenin increased HepG2 cell PI3K activity and decreased insulin receptor substrate (IRS)-2 levels. In sharp contrast to insulin, naringenin did not induce tyrosine phosphorylation of IRS-1. We conclude that naringenin increases LDLr expression in HepG2 cells via PI3K-mediated upregulation of SREBP-1, independent of IRS-1 phosphorylation. Although this pathway may not regulate apoB secretion in primary hepatocytes, PI3K activation by this novel mechanism may explain the insulin-like effects of naringenin in vivo.

  19. Blockade of the phosphatidylinositol-3-kinase-Akt signaling pathway enhances the induction of apoptosis by microtubule-destabilizing agents in tumor cells in which the pathway is constitutively activated.

    PubMed

    Fujiwara, Yusuke; Hosokawa, Yoshihisa; Watanabe, Kazushi; Tanimura, Susumu; Ozaki, Kei-ichi; Kohno, Michiaki

    2007-03-01

    Constitutive activation of the phosphatidylinositol-3-kinase (PI3K)-Akt signaling pathway is associated with the neoplastic phenotype in many human tumor cell types. Given the antiapoptotic role of this pathway, we examined whether its specific blockade might sensitize human tumor cells to the induction of apoptosis by various anticancer drugs. Although specific blockade of the PI3K-Akt pathway alone with inhibitors such as LY294002 did not induce cell death, it resulted in marked and selective enhancement of the induction of apoptosis by microtubule-destabilizing agents such as vincristine. This effect was apparent only in tumor cells in which the PI3K-Akt pathway is constitutively activated. Blockade of the PI3K-Akt pathway induced the activation of glycogen synthase kinase-3beta, which phosphorylates microtubule-associated proteins such as tau and thereby reduces their ability to bind and stabilize microtubules. The consequent destabilization of microtubules induced by the inhibition of PI3K-Akt signaling appeared to increase their sensitivity to low concentrations of microtubule-destabilizing agents that alone do not lead to the disruption of cytoplasmic microtubules in tumor cells. Such a synergistic effect on microtubule integrity was not apparent for stable microtubules in the neurites of neuronal cells. These results suggest that the administration of a combination of a PI3K-Akt pathway inhibitor and a microtubule-destabilizing agent is a potential chemotherapeutic strategy for the treatment of tumor cells in which this signaling pathway is constitutively activated.

  20. Drosophila Mtm and class II PI3K coregulate a PI(3)P pool with cortical and endolysosomal functions.

    PubMed

    Velichkova, Michaella; Juan, Joe; Kadandale, Pavan; Jean, Steve; Ribeiro, Inês; Raman, Vignesh; Stefan, Chris; Kiger, Amy A

    2010-08-09

    Reversible phosphoinositide phosphorylation provides a dynamic membrane code that balances opposing cell functions. However, in vivo regulatory relationships between specific kinases, phosphatases, and phosphoinositide subpools are not clear. We identified myotubularin (mtm), a Drosophila melanogaster MTM1/MTMR2 phosphoinositide phosphatase, as necessary and sufficient for immune cell protrusion formation and recruitment to wounds. Mtm-mediated turnover of endosomal phosphatidylinositol 3-phosphate (PI(3)P) pools generated by both class II and III phosphatidylinositol 3-kinases (Pi3K68D and Vps34, respectively) is needed to down-regulate membrane influx, promote efflux, and maintain endolysosomal homeostasis. Endocytosis, but not endolysosomal size, contributes to cortical remodeling by mtm function. We propose that Mtm-dependent regulation of an endosomal PI(3)P pool has separable consequences for endolysosomal homeostasis and cortical remodeling. Pi3K68D depletion (but not Vps34) rescues protrusion and distribution defects in mtm-deficient immune cells and restores functions in other tissues essential for viability. The broad interactions between mtm and class II Pi3K68D suggest a novel strategy for rebalancing PI(3)P-mediated cell functions in MTM-related human disease.

  1. Drosophila Mtm and class II PI3K coregulate a PI(3)P pool with cortical and endolysosomal functions

    PubMed Central

    Velichkova, Michaella; Juan, Joe; Kadandale, Pavan; Jean, Steve; Ribeiro, Inês; Raman, Vignesh; Stefan, Chris

    2010-01-01

    Reversible phosphoinositide phosphorylation provides a dynamic membrane code that balances opposing cell functions. However, in vivo regulatory relationships between specific kinases, phosphatases, and phosphoinositide subpools are not clear. We identified myotubularin (mtm), a Drosophila melanogaster MTM1/MTMR2 phosphoinositide phosphatase, as necessary and sufficient for immune cell protrusion formation and recruitment to wounds. Mtm-mediated turnover of endosomal phosphatidylinositol 3-phosphate (PI(3)P) pools generated by both class II and III phosphatidylinositol 3-kinases (Pi3K68D and Vps34, respectively) is needed to down-regulate membrane influx, promote efflux, and maintain endolysosomal homeostasis. Endocytosis, but not endolysosomal size, contributes to cortical remodeling by mtm function. We propose that Mtm-dependent regulation of an endosomal PI(3)P pool has separable consequences for endolysosomal homeostasis and cortical remodeling. Pi3K68D depletion (but not Vps34) rescues protrusion and distribution defects in mtm-deficient immune cells and restores functions in other tissues essential for viability. The broad interactions between mtm and class II Pi3K68D suggest a novel strategy for rebalancing PI(3)P-mediated cell functions in MTM-related human disease. PMID:20696708

  2. p85 regulatory subunit of PI3K mediates cAMP-PKA and estrogens biological effects on growth and survival.

    PubMed

    Cosentino, C; Di Domenico, M; Porcellini, A; Cuozzo, C; De Gregorio, G; Santillo, M R; Agnese, S; Di Stasio, R; Feliciello, A; Migliaccio, A; Avvedimento, E V

    2007-03-29

    Cyclic adenosine 3'5' monophosphate (cAMP) and protein kinase A (PKA) cooperate with phosphatidylinositol 3' kinase (PI3K) signals in the control of growth and survival. To determine the molecular mechanism(s) involved, we identified and mutagenized a specific serine (residue 83) in p85alpha(PI3K), which is phosphorylated in vivo and in vitro by PKA. Expression of p85alpha(PI3K) mutants (alanine or aspartic substitutions) significantly altered the biological responses of the cells to cAMP. cAMP protection from anoikis was reduced in cells expressing the alanine version p85alpha(PI3K). These cells did not arrest in G1 in the presence of cAMP, whereas cells expressing the aspartic mutant p85D accumulated in G1 even in the absence of cAMP. S phase was still efficiently inhibited by cAMP in cells expressing both mutants. The binding of PI3K to Ras p21 was greatly reduced in cells expressing p85A in the presence or absence of cAMP. Conversely, expression of the aspartic mutant stimulated robustly the binding of PI3K to p21 Ras in the presence of cAMP. Mutation in the Ser 83 inhibited cAMP, but not PDGF stimulation of PI3K. Conversely, the p85D aspartic mutant amplified cAMP stimulation of PI3K activity. Phosphorylation of Ser 83 by cAMP-PKA in p85alpha(PI3K) was also necessary for estrogen signaling as expression of p85A or p85D mutants inhibited or amplified, respectively, the binding of estrogen receptor to p85alpha and AKT phosphorylation induced by estrogens. The data presented indicate that: (1) phosphorylation of Ser 83 in p85alpha(PI3K) is critical for cAMP-PKA induced G1 arrest and survival in mouse 3T3 fibroblasts; (2) this site is necessary for amplification of estrogen signals by cAMP-PKA and related receptors. Finally, these data suggest a general mechanism of PI3K regulation by cAMP, operating in various cell types and under different conditions.

  3. Deciphering Combinations of PI3K/AKT/mTOR Pathway Drugs Augmenting Anti-Angiogenic Efficacy In Vivo

    PubMed Central

    Sasore, Temitope; Kennedy, Breandán

    2014-01-01

    Ocular neovascularization is a common pathology associated with human eye diseases e.g. age-related macular degeneration and proliferative diabetic retinopathy. Blindness represents one of the most feared disabilities and remains a major burden to health-care systems. Current approaches to treat ocular neovascularisation include laser photocoagulation, photodynamic therapy and anti-VEGF therapies: Ranibizumab (Lucentis) and Aflibercept (Eylea). However, high clinical costs, frequent intraocular injections, and increased risk of infections are challenges related with these standards of care. Thus, there is a clinical need to develop more effective drugs that overcome these challenges. Here, we focus on an alternative approach by quantifying the in vivo anti-angiogenic efficacy of combinations of phosphatidylinositol-3-kinase (PI3K) pathway inhibitors. The PI3K/AKT/mTOR pathway is a complex signalling pathway involved in crucial cellular functions such as cell proliferation, migration and angiogenesis. RT-PCR confirms the expression of PI3K target genes (pik3ca, pik3r1, mtor and akt1) in zebrafish trunks from 6 hours post fertilisation (hpf) and in eyes from 2 days post fertilisation (dpf). Using both the zebrafish intersegmental vessel and hyaloid vessel assays to measure the in vivo anti-angiogenic efficacy of PI3K/Akt/mTOR pathway inhibitors, we identified 5 µM combinations of i) NVP-BEZ235 (dual PI3K-mTOR inhibitor) + PI-103 (dual PI3K-mTOR inhibitor); or ii) LY-294002 (pan-PI3K inhibitor) + NVP-BEZ235; or iii) NVP-BEZ235 + rapamycin (mTOR inhibitor); or iv) LY-294002 + rapamycin as the most anti-angiogenic. Treatment of developing larvae from 2–5 dpf with 5 µM NVP-BEZ235 plus PI-103 resulted in an essentially intact ocular morphology and visual behaviour, whereas other combinations severely disrupted the developing retinal morphology and visual function. In human ARPE19 retinal pigment epithelium cells, however, no significant difference in cell number was

  4. Regulation of PI3K effector signalling in cancer by the phosphoinositide phosphatases

    PubMed Central

    Rodgers, Samuel J.; Ferguson, Daniel T.; Mitchell, Christina A.

    2017-01-01

    Class I phosphoinositide 3-kinase (PI3K) generates phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) at the plasma membrane in response to growth factors, activating a signalling cascade that regulates many cellular functions including cell growth, proliferation, survival, migration and metabolism. The PI3K pathway is commonly dysregulated in human cancer, and drives tumorigenesis by promoting aberrant cell growth and transformation. PtdIns(3,4,5)P3 facilitates the activation of many pleckstrin homology (PH) domain-containing proteins including the serine/threonine kinase AKT. There are three AKT isoforms that are frequently hyperactivated in cancer through mutation, amplification or dysregulation of upstream regulatory proteins. AKT isoforms have converging and opposing functions in tumorigenesis. PtdIns(3,4,5)P3 signalling is degraded and terminated by phosphoinositide phosphatases such as phosphatase and tensin homologue (PTEN), proline-rich inositol polyphosphate 5-phosphatase (PIPP) (INPP5J) and inositol polyphosphate 4-phosphatase type II (INPP4B). PtdIns(3,4,5)P3 is rapidly hydrolysed by PIPP to generate phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2), which is further hydrolysed by INPP4B to form phosphatidylinositol 3-phosphate (PtdIns3P). PtdIns(3,4)P2 and PtdIns3P are also important signalling molecules; PtdIns(3,4)P2 together with PtdIns(3,4,5)P3 are required for maximal AKT activation and PtdIns3P activates PI3K-dependent serum and glucocorticoid-regulated kinase (SGK3) signalling. Loss of Pten, Pipp or Inpp4b expression or function promotes tumour growth in murine cancer models through enhanced AKT isoform-specific signalling. INPP4B inhibits PtdIns(3,4)P2-mediated AKT activation in breast and prostate cancer; however, INPP4B expression is increased in acute myeloid leukaemia (AML), melanoma and colon cancer where it paradoxically promotes cell proliferation, transformation and/or drug resistance. This review will discuss how PTEN, PIPP

  5. A Single Oral Dose of Geranylgeranylacetone Upregulates Vascular Endothelial Growth Factor and Protects against Kainic Acid-Induced Neuronal Cell Death: Involvement of the Phosphatidylinositol-3 Kinase/Akt Pathway.

    PubMed

    Kawasaki, Yukari; Fujiki, Minoru; Uchida, Susumu; Morishige, Masaki; Momii, Yasutomo; Ishii, Keisuke

    2017-01-01

    Previous studies demonstrated the cytoprotective effect of geranylgeranylacetone (GGA), a heat shock protein inducer, against ischemic insult or kainic acid (KA)-induced neuronal cell death. Phosphatidylinositol-3 kinase (PI3K)/Akt is thought to be an important factor that mediates neuroprotection. However, the signaling pathways in the brain in vivo after oral GGA administration remain unclear. We measured and compared hippocampal neuron density to investigate the effect of GGA on KA-induced cell death in rats. We evaluated the effects of pretreatment with wortmannin (Wort), a specific PI3K inhibitor, on GGA-induced neuroprotection against KA-induced cell death. To clarify the relationship between PI3K/Akt activation and neuroprotection, we used immunoblot analysis to determine the amounts of p-Akt and vascular endothelial growth factor (VEGF) proteins present after GGA administration with or without Wort treatment. Neuroprotective effects of GGA (pretreatment with a single oral dose of GGA, 800 mg/kg, 48 h before KA injection) were prevented by Wort pretreatment, which indicates that the selective PI3K/Akt pathway may mediate the GGA-dependent protection. Oral GGA-induced p-Akt and VEGF, and GGA pretreatment enhanced KA-induced VEGF, both of which were prevented by Wort pretreatment. These results suggest that a single oral dose of GGA induces p-Akt and that GGA plays an important role in neuroprotection against KA-induced neuronal cell death through VEGF induction. © 2017 S. Karger AG, Basel.

  6. PI3K pan-inhibition impairs more efficiently proliferation and survival of T-cell acute lymphoblastic leukemia cell lines when compared to isoform-selective PI3K inhibitors

    PubMed Central

    Spartà, Antonino Maria; Chiarini, Francesca; Buontempo, Francesca; Evangelisti, Camilla; Evangelisti, Cecilia; Orsini, Ester; McCubrey, James A.; Martelli, Alberto Maria

    2015-01-01

    Class I phosphatidylinositol 3-kinases (PI3Ks) are frequently activated in T-cell acute lymphoblastic leukemia (T-ALL), mainly due to the loss of PTEN function. Therefore, targeting PI3Ks is a promising innovative approach for T-ALL treatment, however at present no definitive evidence indicated which is the better therapeutic strategy between pan or selective isoform inhibition, as all the four catalytic subunits might participate in leukemogenesis. Here, we demonstrated that in both PTEN deleted and PTEN non deleted T-ALL cell lines, PI3K pan-inhibition exerted the highest cytotoxic effects when compared to both selective isoform inhibition or dual p110γ/δ inhibition. Intriguingly, the dual p110γ/δ inhibitor IPI-145 was effective in Loucy cells, which are representative of early T-precursor (ETP)-ALL, a T-ALL subtype associated with a poor outcome. PTEN gene deletion did not confer a peculiar reliance of T-ALL cells on PI3K activity for their proliferation/survival, as PTEN was inactivated in PTEN non deleted cells, due to posttranslational mechanisms. PI3K pan-inhibition suppressed Akt activation and induced caspase-independent apoptosis. We further demonstrated that in some T-ALL cell lines, autophagy could exert a protective role against PI3K inhibition. Our findings strongly support clinical application of class I PI3K pan-inhibitors in T-ALL treatment, with the possible exception of ETP-ALL cases. PMID:25871383

  7. Sevoflurane Postconditioning Protects Rat Hearts against Ischemia-Reperfusion Injury via the Activation of PI3K/AKT/mTOR Signaling

    PubMed Central

    Zhang, Jing; Wang, Chen; Yu, Shuchun; Luo, Zhenzhong; Chen, Yong; Liu, Qin; Hua, Fuzhou; Xu, Guohai; Yu, Peng

    2014-01-01

    Phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway plays a key role in myocardial ischemia-reperfusion (I/R) injury. Mammalian target of rapamycin (mTOR), a downstream target of PI3K/AKT signaling, is necessary and sufficient to protect the heart from I/R injury. Inhaled anesthetic sevoflurane is widely used in cardiac surgeries because its induction and recovery are faster and smoother than other inhaled anesthetics. Sevoflurane proved capable of inducing postconditioning effects in the myocardium. However, the underlying molecular mechanisms for sevoflurane-induced postconditioning (SPC) were largely unclear. In the present study, we demonstrated that SPC protects myocardium from I/R injury with narrowed cardiac infarct focus, increased ATP content, and decreased cardiomyocyte apoptosis, which are mainly due to the activation of PI3K/AKT/mTOR signaling and the protection of mitochondrial energy metabolism. Application of dactolisib (BEZ235), a PI3K/mTOR dual inhibitor, abolishes the up-regulation of pho-AKT, pho-GSK, pho-mTOR, and pho-p70s6k induced by SPC, hence abrogating the anti-apoptotic effect of sevoflurane and reducing SPC-mediated protection of heart from I/R injury. As such, this study proved that PI3K/AKT/mTOR pathway plays an important role in SPC induced cardiac protection against I/R injury. PMID:25471136

  8. The Role of the PI3K Pathway in the Regeneration of the Damaged Brain by Neural Stem Cells after Cerebral Infarction.

    PubMed

    Koh, Seong Ho; Lo, Eng H

    2015-10-01

    Neurologic deficits resulting from stroke remain largely intractable, which has prompted thousands of studies aimed at developing methods for treating these neurologic sequelae. Endogenous neurogenesis is also known to occur after brain damage, including that due to cerebral infarction. Focusing on this process may provide a solution for treating neurologic deficits caused by cerebral infarction. The phosphatidylinositol-3-kinase (PI3K) pathway is known to play important roles in cell survival, and many studies have focused on use of the PI3K pathway to treat brain injury after stroke. Furthermore, since the PI3K pathway may also play key roles in the physiology of neural stem cells (NSCs), eliciting the appropriate activation of the PI3K pathway in NSCs may help to improve the sequelae of cerebral infarction. This review describes the PI3K pathway, its roles in the brain and NSCs after cerebral infarction, and the therapeutic possibility of activating the pathway to improve neurologic deficits after cerebral infarction.

  9. PI3K/Akt signaling mediated apoptosis blockage and viral gene expression in oral epithelial cells during herpes simplex virus infection.

    PubMed

    Hsu, Mei-Ju; Wu, Ching-Yi; Chiang, Hsiao-Han; Lai, Yu-Lin; Hung, Shan-Ling

    2010-10-01

    Phosphatidylinositol 3-kinases (PI3Ks) function in the anti-apoptotic pathway, and are commonly exploited by various viruses to accomplish the viral life cycle. This study examined the role of the PI3K pathway in human oral epithelial cells following herpes simplex virus type 1 (HSV-1) infection. The results showed that HSV-1 induced the phosphorylation of Akt and glycogen synthase kinase 3 (GSK-3). Phosphorylation of Akt, but not GSK-3, induced by HSV-1 was PI3K-dependent. The expression of HSV-1 immediate-early genes may be involved in the initial phosphorylation of Akt and GSK-3. Inhibition of HSV-1-induced PI3K activity increased DNA fragmentation and cleavage of poly ADP-ribose polymerase (PARP), caspase 3 and caspase 7 compared with infected alone. Inhibition of PI3K attenuated the expression of HSV-1-infected cell protein 0 (ICP0), but not thymidine kinase (TK) and viral replication. Collectively, these data suggested that, in oral epithelial cells, the HSV-1-induced PI3K/Akt activation was involved in the regulation of apoptosis blockage and viral gene expression.

  10. Fibroblast growth factor 2 induces E-cadherin down-regulation via PI3K/Akt/mTOR and MAPK/ERK signaling in ovarian cancer cells.

    PubMed

    Lau, Man-Tat; So, Wai-Kin; Leung, Peter C K

    2013-01-01

    Fibroblast growth factor 2 (FGF2) is produced by ovarian cancer cells and it has been suggested to play an important role in tumor progression. In this study, we report that FGF2 treatment down-regulated E-cadherin by up-regulating its transcriptional repressors, Slug and ZEB1, in human ovarian cancer cells. The pharmacological inhibition of phosphatidylinositol-3-kinase (PI3K), mammalian target of rapamycin (mTOR), and MEK suggests that both PI3K/Akt/mTOR and MAPK/ERK signaling are required for FGF2-induced E-cadherin down-regulation. Moreover, FGF2 up-regulated Slug and ZEB1 expression via the PI3K/Akt/mTOR and MAPK/ERK signaling pathways, respectively. Finally, FGF2-induced cell invasion was abolished by the inhibition of the PI3K/Akt/mTOR and MAPK/ERK pathways, and the forced expression of E-cadherin diminished the intrinsic invasiveness of ovarian cancer cells as well as the FGF2-induced cell invasion. This study demonstrates a novel mechanism in which FGF2 down-regulates E-cadherin expression through the activation of PI3K/Akt/mTOR and MAPK/ERK signaling, and the up-regulation of Slug and ZEB1 in human ovarian cancer cells.

  11. The Role of the PI3K Pathway in the Regeneration of the Damaged Brain by Neural Stem Cells after Cerebral Infarction

    PubMed Central

    Lo, Eng H.

    2015-01-01

    Neurologic deficits resulting from stroke remain largely intractable, which has prompted thousands of studies aimed at developing methods for treating these neurologic sequelae. Endogenous neurogenesis is also known to occur after brain damage, including that due to cerebral infarction. Focusing on this process may provide a solution for treating neurologic deficits caused by cerebral infarction. The phosphatidylinositol-3-kinase (PI3K) pathway is known to play important roles in cell survival, and many studies have focused on use of the PI3K pathway to treat brain injury after stroke. Furthermore, since the PI3K pathway may also play key roles in the physiology of neural stem cells (NSCs), eliciting the appropriate activation of the PI3K pathway in NSCs may help to improve the sequelae of cerebral infarction. This review describes the PI3K pathway, its roles in the brain and NSCs after cerebral infarction, and the therapeutic possibility of activating the pathway to improve neurologic deficits after cerebral infarction. PMID:26320845

  12. PI3K p110δ uniquely promotes gain-of-function Shp2-induced GM-CSF hypersensitivity in a model of JMML.

    PubMed

    Goodwin, Charles B; Li, Xing Jun; Mali, Raghuveer S; Chan, Gordon; Kang, Michelle; Liu, Ziyue; Vanhaesebroeck, Bart; Neel, Benjamin G; Loh, Mignon L; Lannutti, Brian J; Kapur, Reuben; Chan, Rebecca J

    2014-05-01

    Although hyperactivation of the Ras-Erk signaling pathway is known to underlie the pathogenesis of juvenile myelomonocytic leukemia (JMML), a fatal childhood disease, the PI3K-Akt signaling pathway is also dysregulated in this disease. Using genetic models, we demonstrate that inactivation of phosphatidylinositol-3-kinase (PI3K) catalytic subunit p110δ, but not PI3K p110α, corrects gain-of-function (GOF) Shp2-induced granulocyte macrophage-colony-stimulating factor (GM-CSF) hypersensitivity, Akt and Erk hyperactivation, and skewed hematopoietic progenitor distribution. Likewise, potent p110δ-specific inhibitors curtail the proliferation of GOF Shp2-expressing hematopoietic cells and cooperate with mitogen-activated or extracellular signal-regulated protein kinase kinase (MEK) inhibition to reduce proliferation further and maximally block Erk and Akt activation. Furthermore, the PI3K p110δ-specific inhibitor, idelalisib, also demonstrates activity against primary leukemia cells from individuals with JMML. These findings suggest that selective inhibition of the PI3K catalytic subunit p110δ could provide an innovative approach for treatment of JMML, with the potential for limiting toxicity resulting from the hematopoietic-restricted expression of p110δ.

  13. Expression of the RAE-1 Family of Stimulatory NK-Cell Ligands Requires Activation of the PI3K Pathway during Viral Infection and Transformation

    PubMed Central

    Tokuyama, Maria; Lorin, Clarisse; Delebecque, Frederic; Jung, Heiyoun; Raulet, David H.; Coscoy, Laurent

    2011-01-01

    Natural killer (NK) cells are lymphocytes that play a major role in the elimination of virally-infected cells and tumor cells. NK cells recognize and target abnormal cells through activation of stimulatory receptors such as NKG2D. NKG2D ligands are self-proteins, which are absent or expressed at low levels on healthy cells but are induced upon cellular stress, transformation, or viral infection. The exact molecular mechanisms driving expression of these ligands remain poorly understood. Here we show that murine cytomegalovirus (MCMV) infection activates the phosphatidylinositol-3-kinase (PI3K) pathway and that this activation is required for the induction of the RAE-1 family of mouse NKG2D ligands. Among the multiple PI3K catalytic subunits, inhibition of the p110α catalytic subunit blocks this induction. Similarly, inhibition of p110α PI3K reduces cell surface expression of RAE-1 on transformed cells. Many viruses manipulate the PI3K pathway, and tumors frequently mutate the p110α oncogene. Thus, our findings suggest that dysregulation of the PI3K pathway is an important signal to induce expression of RAE-1, and this may represent a commonality among various types of cellular stresses that result in the induction of NKG2D ligands. PMID:21966273

  14. Suppression of Virulent Porcine Epidemic Diarrhea Virus Proliferation by the PI3K/Akt/GSK-3α/β Pathway

    PubMed Central

    Kong, Ning; Wu, Yongguang; Meng, Qiong; Wang, Zhongze; Zuo, Yewen; Pan, Xi; Tong, Wu; Zheng, Hao; Li, Guoxin; Yang, Shen; Yu, Hai; Zhou, En-min; Shan, Tongling; Tong, Guangzhi

    2016-01-01

    Porcine epidemic diarrhea virus (PEDV) has recently caused high mortality in suckling piglets with subsequent large economic losses to the swine industry. Many intracellular signaling pathways, including the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, are activated by viral infection. The PI3K/Akt pathway is an important cellular pathway that has been shown to be required for virus replication. In the present study, we found that the PEDV JS-2013 strain activated Akt in Vero cells at early (5–15 min) and late stages (8–10 h) of infection. Inhibiting PI3K, an upstream activator of Akt, enhanced PEDV replication. Inhibiting GSK-3α/β, one of the downstream effectors of PI3K/Akt pathway and regulated by Akt during PEDV infected Vero cells, also enhanced PEDV replication. Collectively, our data suggest that PI3K/Akt/GSK-3α/β signaling pathway is activated by PEDV and functions in inhibiting PEDV replication. PMID:27560518

  15. Andrographolide inhibits hypoxia-inducible factor-1 through phosphatidylinositol 3-kinase/AKT pathway and suppresses breast cancer growth

    PubMed Central

    Li, Jie; Zhang, Chao; Jiang, Hongchuan; Cheng, Jiao

    2015-01-01

    Hypoxia-inducible factor-1 (HIF-1) is a master regulator of the transcriptional response to hypoxia. HIF-1α is one of the most compelling anticancer targets. Andrographolide (Andro) was newly identified to inhibit HIF-1 in T47D cells (a half maximal effective concentration [EC50] of 1.03×10−7 mol/L), by a dual-luciferase reporter assay. It suppressed HIF-1α protein and gene accumulation, which was dependent on the inhibition of upstream phosphatidylinositol 3-kinase (PI3K)/AKT pathway. It also abrogated the expression of HIF-1 target vascular endothelial growth factor (VEGF) gene and protein. Further, Andro inhibited T47D and MDA-MB-231 cell proliferation and colony formation. In addition, it exhibited significant in vivo efficacy and antitumor potential against the MDA-MB-231 xenograft in nude mice. In conclusion, these results highlighted the potential effects of Andro, which inhibits HIF-1, and hence may be developed as an antitumor agent for breast cancer therapy in future. PMID:25709476

  16. Secretin Receptor Promotes the Proliferation of Endocrine Tumor Cells Via the PI3K/AKT Pathway

    PubMed Central

    Lee, Misu; Waser, Beatrice; Reubi, Jean-Claude

    2012-01-01

    The secretin receptor (SR), a G protein-coupled receptor, mediates the effects of the gastrointestinal hormone secretin on digestion and water homeostasis. Recently, high SR expression has been observed in pancreatic ductal adenocarcinomas, cholangiocellular carcinomas, gastrinomas, and bronchopulmonary carcinoid tumors. Receptor overexpression associates with enhanced secretin-mediated signaling, but whether this molecule plays an independent role in tumorigenesis is currently unknown. We recently discovered that pheochromocytomas developing in rats affected by the MENX (multiple endocrine neoplasia-like) syndrome express at very high-level Sctr, encoding SR. We here report that SR are also highly abundant on the membranes of rat adrenal and extraadrenal pheochromocytoma, starting from early stages of tumor development, and are functional. PC12 cells, the best characterized in vitro pheochromocytoma model, also express Sctr at high level. Thus, we used them as model to study the role of SR in neoplastic transformation. Small interfering RNA-mediated knockdown of Sctr decreases PC12 cells proliferation and increases p27 levels. The proproliferative effect of SR in PC12 cells is mediated, in part, by the phosphatidylinositol 3 kinase (PI3K)/serine-threonine protein kinase (AKT) pathway. Transfection of Sctr in Y1 adrenocortical carcinoma cells, expressing low endogenous levels of Sctr, stimulates cell proliferation also, in part, via the PI3K/AKT signaling cascade. Because of the link between SR and PI3K/AKT signaling, tumor cells expressing high levels of the receptor (MENX-associated primary pheochromocytoma and NCI-H727 human bronchopulmonary carcinoid cells) respond well and in a SR-dependent manner to PI3K inhibitors, such as NVP-BEZ235. The association between SR levels and response to PI3K inhibition might open new avenues for the treatment of tumors overexpressing this receptor. PMID:22692904

  17. Treadmill exercise improves motor and memory functions in cerebral palsy rats through activation of PI3K-Akt pathway.

    PubMed

    Jung, Sun-Young; Kim, Dae-Young

    2017-04-01

    Cerebral palsy (CP) is a chronic disorder characterized by physical disability and disruption of brain function. We evaluated the effects of treadmill exercise on motor and memory functions in relation with phosphatidylinositol 3-kinase (PI3K)-Akt pathway using CP rat model. Rota-rod test, step-down avoidance task, 5-bromo-2'-deoxyuridine (BrdU) immunohistochemistry, and western blot for synapsin I, postsynaptic density-95 (PSD-95), PI3K, Akt, and glycogen synthase kinase-3β (GSK-3β) were performed. CP was induced by maternal lipopolysaccharide (LPS)-injection with sensorimotor restriction. Five weeks after birth, the rats in the exercise groups were made to run on the treadmill for 30 min per one day, 5 times a week, during 4 weeks. Motor and memory functions were impaired in the LPS-induced CP rats and tread-mill exercise increased motor and memory functions in the CP rats. Cell proliferation in the hippocampus was suppressed in the LPS-induced CP rats and treadmill exercise increased hippocampal cell proliferation in the CP rats. Expressions of synapsin I, PSD-95, phosphorylated (p)-PI3K, and p-Akt were decreased in the LPS-induced CP rats and treadmill exercise enhanced the expressions of synapsin I, PSD-95, p-PI3K, and p-Akt in the CP rats. GSK-3β expression was increased in the LPS-induced CP rats and treadmill exercise suppressed GSK-3β expression in the CP rats. The present results suggest that treadmill exercise might improve motor and memory functions through activation of PI3K-Akt pathway.

  18. Treadmill exercise improves motor and memory functions in cerebral palsy rats through activation of PI3K-Akt pathway

    PubMed Central

    Jung, Sun-Young; Kim, Dae-Young

    2017-01-01

    Cerebral palsy (CP) is a chronic disorder characterized by physical disability and disruption of brain function. We evaluated the effects of treadmill exercise on motor and memory functions in relation with phosphatidylinositol 3-kinase (PI3K)-Akt pathway using CP rat model. Rota-rod test, step-down avoidance task, 5-bromo-2′-deoxyuridine (BrdU) immunohistochemistry, and western blot for synapsin I, postsynaptic density-95 (PSD-95), PI3K, Akt, and glycogen synthase kinase-3β (GSK-3β) were performed. CP was induced by maternal lipopolysaccharide (LPS)-injection with sensorimotor restriction. Five weeks after birth, the rats in the exercise groups were made to run on the treadmill for 30 min per one day, 5 times a week, during 4 weeks. Motor and memory functions were impaired in the LPS-induced CP rats and tread-mill exercise increased motor and memory functions in the CP rats. Cell proliferation in the hippocampus was suppressed in the LPS-induced CP rats and treadmill exercise increased hippocampal cell proliferation in the CP rats. Expressions of synapsin I, PSD-95, phosphorylated (p)-PI3K, and p-Akt were decreased in the LPS-induced CP rats and treadmill exercise enhanced the expressions of synapsin I, PSD-95, p-PI3K, and p-Akt in the CP rats. GSK-3β expression was increased in the LPS-induced CP rats and treadmill exercise suppressed GSK-3β expression in the CP rats. The present results suggest that treadmill exercise might improve motor and memory functions through activation of PI3K-Akt pathway. PMID:28503524

  19. Paclitaxel resistance in MCF-7/PTX cells is reversed by paeonol through suppression of the SET/phosphatidylinositol 3-kinase/Akt pathway.

    PubMed

    Zhang, Weipeng; Cai, Jiangxia; Chen, Siying; Zheng, Xiaowei; Hu, Sasa; Dong, Weihua; Lu, Jun; Xing, Jianfeng; Dong, Yalin

    2015-07-01

    Breast cancer is one of the most prevalent types of malignant tumor. Paclitaxel is widely used in the treatment of breast cancer; however, the major problem contributing to the failure of chemotherapy in breast cancer is the development of drug resistance. Therefore, it is necessary to identify novel therapeutic targets and reversal agents for breast cancer. In the present study, the protein expression levels of SET, protein phosphatase 2A (PP2A) and phosphatidylinositol 3-kinase (PI3K)/Akt pathway were determined in MCF-7/PTX human breast carcinoma paclitaxel-resistant cells using western blot analysis. Small interference RNAs (siRNAs) were used to knock down the gene expression of SET in MCF-7/PTX cells and the cell viability was assessed following treatment with paclitaxel, using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assays and flow cytometry. In addition, western blot analysis was used to determined PI3K/Akt pathway activity following SET knockdown. Furthermore, the reversal effects of paeonol on paclitaxel, and its underlying mechanisms of action, were investigated using western blot analysis and reverse transcription-quantitative polymerase chain reaction. The results demonstrated that increased levels of SET and PI3K/Akt pathway proteins were present in the MCF-7/PTX cells, compared with normal MCF-7 cells. Knockdown of SET significantly sensitized MCF-7/PTX cells to paclitaxel and induced cell apoptosis. In addition, the expression levels of the adenosine triphosphate binding cassette (ABC) transporter proteins were significantly reduced in the MCF-7/PTX cells compared with the normal MCF-7 cells. SET-induced paclitaxel resistance was found to be associated with the activation of the PI3K/Akt pathway. Paeonol significantly reduced the mRNA and protein expression levels of SET in the MCF-7/PTX cells. Furthermore, paeonol significantly sensitized the MCF-7/PTX to paclitaxel via regulation of ABC transporters, B cell lymphoma-2 (Bcl-2

  20. Type II cGMP-dependent protein kinase inhibits epidermal growth factor-induced phosphatidylinositol-3-kinase/Akt signal transduction in gastric cancer cells.

    PubMed

    Wu, Min; Chen, Yongchang; Jiang, Lu; Li, Yueying; Lan, Ting; Wang, Ying; Qian, Hai

    2013-12-01

    Our previous study revealed that Type II cGMP-dependent protein kinase (PKG II) inhibits epidermal growth factor (EGF)-induced MAPK/ERK and MAPK/JNK-mediated signal transduction through the inhibition of the phosphorylation/activation of the EGF receptor (EGFR). As EGFR also mediates several other signal transduction pathways besides MAPK-mediated pathways, the present study was designed to investigate whether PKG II was able to inhibit EGF/EGFR-induced phosphatidylinositol-3-kinase (PI3K)/Akt-mediated signal transduction. The AGS human gastric cancer cell line was infected with adenoviral constructs encoding a cDNA of PKG II (Ad-PKG II) to increase the expression of PKG II, and treated with 8-pCPT-cGMP to activate the enzyme. Western blotting was used to detect the phosphorylation/activation of the key components of the signal transduction pathway, including EGFR, PI3K, Akt, mTOR and NF-κB. The levels of apoptosis-related proteins, including Bax, Bcl-2, caspase 9 and DNA fragment factor (DFF), were also determined by western blotting. Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling staining was used to detect the apoptosis of the AGS cells. The results revealed that EGF treatment increased the phosphorylation (activation) of EGFR, PI3K, Akt and mTOR, and increased the nuclear localization (activation) of NF-κB. EGF treatment also reduced the apoptosis of the AGS cells and increased the expression of the anti-apoptotic protein, Bcl-2, but had no effect on the expression of the pro-apoptotic protein, Bax, and did not alter the levels of caspase 9 and DFF. Increasing the PKG II activity of AGS cells by infecting them with Ad-PKG II and stimulating them with 8-pCPT-cGMP inhibited the EGF-induced activation of EGFR, PI3K, Akt, mTOR and NF-κB; caused an increase in caspase 9 breakdown (activation) and DFF levels; and reversed the anti-apoptotic effect of EGF. The results suggest that PKG II may also inhibit EGF-induced signal transduction of PI3

  1. Role of the phosphatidylinositol 3-kinase/Akt and mTOR/P70S6-kinase pathways in the proliferation and apoptosis in multiple myeloma.

    PubMed

    Pene, Frédéric; Claessens, Yann-Erick; Muller, Odile; Viguié, Franck; Mayeux, Patrick; Dreyfus, François; Lacombe, Catherine; Bouscary, Didier

    2002-09-26

    Multiple myeloma (MM) is a plasma cell malignancy preliminary localized in the bone marrow and characterized by its capacity to disseminate. IL-6 and IGF-1 have been shown to mediate proliferative and anti-apoptotic signals in plasmocytes. However, in primary plasma-cell leukemia (PCL) and in end-stage aggressive extramedullar disease, the cytokine requirement for both effects may be not mandatory. This suggests that constitutive activation of signaling pathways occurs. One of the signaling pathways whose deregulation may play an oncogenic role in MM is the phosphatidylinositol 3-kinase (PI 3-K) pathway. In human growth factor-independent MM cell lines OPM2 and RPMI8226, we show that the PI 3-K inhibitors LY294002 and Wortmannin strongly inhibited cell proliferation, whereas inhibition of the mammalian Target Of Rapamycin (mTOR)/P70-S6-kinase (P70(S6K)) pathway with rapamycin or of the Mitogen-Activated Protein Kinase (MAPK) pathway with PD98059 had minimal effect on proliferation. In both cell lines, constitutive activation of the PI 3-K/Akt/FKHRL-1, mTOR/P70(S6K) and MAPK pathways was detected. LY294002 inhibited phosphorylation of Akt, FKHRL-1 and P70(S6K) but had no effect on ERK1/2 phosphorylation, indicating that the PI 3-K and MAPK pathways are independent. IGF-1 but not IL-6 increased phosphorylation of Akt, FKHRL-1 and P70(S6K). Purified plasmocytes from four patients with MM and two patients with primary PCL were studied. In three of them including the two patients with PCL, constitutive phosphorylation of Akt, FKHRL-1 and P70(S6K) was present, inhibited by LY294002 and enhanced by IGF-1. In these patients with constitutive Akt activation, normal PTEN expression was detected. PI 3-K inhibition induced caspase-dependent apoptosis as confirmed by inhibition with the large spectrum caspase inhibitor Z-VAD-FMK and cleavage of pro-caspase-3. Both cell lines spontaneously expressed Skp2 and cyclin D1 proteins at high levels but no p27(Kip1) protein. In the

  2. Efficacy of phosphatidylinositol-3 kinase inhibitors with diverse isoform selectivity profiles for inhibiting the survival of chronic lymphocytic leukemia cells.

    PubMed

    Göckeritz, Elisa; Kerwien, Susan; Baumann, Michael; Wigger, Marion; Vondey, Verena; Neumann, Lars; Landwehr, Thomas; Wendtner, Clemens M; Klein, Christian; Liu, Ningshu; Hallek, Michael; Frenzel, Lukas P; Krause, Günter

    2015-11-01

    Pharmacological inhibition of phosphatiylinositide-3-kinase (PI3K)-mediated signaling holds great promise for treating chronic lymphocytic leukemia (CLL). Therefore we assessed three structurally related PI3K inhibitors targeting the PI3K-δ isoform for their ability to inhibit the survival of freshly isolated CLL cells. The purely PI3K-δ-selective inhibitor idelalisib was compared to copanlisib (BAY 80-6946) and duvelisib (IPI-145), with isoform target profiles that additionally include PI3K-α or PI3K-γ, respectively. The concentrations leading to half-maximal reduction of the survival of CLL cells were more than ten-fold lower for copanlisib than for idelalisib and duvelisib. At concentrations reflecting the biological availability of the different inhibitors, high levels of apoptotic response among CLL samples were attained more consistently with copanlisib than with idelalisib. Copanlisib selectively reduced the survival of CLL cells compared to T cells and to B cells from healthy donors. In addition copanlisib and duvelisib impaired the migration of CLL cells towards CXCL12 to a greater extent than equimolar idelalisib. Similarly copanlisib and duvelisib reduced the survival of CLL cells in co-cultures with the bone marrow stroma cell line HS-5 more strongly than idelalisib. Survival inhibition by copanlisib and idelalisib was enhanced by the monoclonal CD20 antibodies rituximab and obinutuzumab (GA101), while antibody-dependent cellular cytotoxicity mediated by alemtuzumab and peripheral blood mononuclear cells was not substantially impaired by both PI3K inhibitors for the CLL-derived JVM-3 cell line as target cells. Taken together, targeting the α- and δ- p110 isoforms with copanlisib may be a useful strategy for the treatment of CLL and warrants further clinical investigation.

  3. The effect of Liuwei Dihuang decoction on PI3K/Akt signaling pathway in liver of type 2 diabetes mellitus (T2DM) rats with insulin resistance.

    PubMed

    Dai, Bing; Wu, Qinxuan; Zeng, Chengxi; Zhang, Jiani; Cao, Luting; Xiao, Zizeng; Yang, Menglin

    2016-11-04

    Liuwei Dihaung decoction (LWDHT) is a well-known classic traditional Chinese medicine formula, consists of six herbs including Rehmannia glutinosa Libosch.(family: Scrophulariaceae), Cornus officinalis Sieb.(family: Cornaceae), Dioscorea opposite Thunb.(family: Dioscoreaceae), Alisma orientale(G. Samuelsson) Juz (family: Alismataceae), Poria cocos (Schw.) Wolf (family: Polyporaceae) and Paeonia suffruticosa Andrews (family: Paeoniaceae). It has been used in the treatment of many types of diseases with signs of deficiency of Yin in the kidneys in China clinically. This study is aimed at investigating the effect of Liuwei dihuang decoction on PI3K/Akt signaling pathway in liver of T2DM rats with insulin resistance. T2DM model was induced in male Sprague-Dawley (SD) rats by high sugar and high fat diets combined with small dose of streptozocin (STZ) injection. The successful T2DM rats were randomly allocated three group--vehicle group, positive control group and Liuwei Dihuang decoction group. After 12-weeks treatment with distilled water, rosiglitazone and LWDHT by intragastric administration respectively, the rats were put to death in batches. The variance of fasting blood glucose (FBG) and fasting insulin (FINS) in serum were determined, the pathological changes of each rats' liver were observed by hematoxylin-eosin (HE) staining, the expression of insulin receptor substrate 2(IRS2), phosphatidylinositol 3-kinase (PI3K) and protein kinas B (Akt) involving the canonical PI3K/Akt signaling pathway were detected by Real-time fluorescent quantitative PCR (RT-PCR), and the expression level of IRS2, PI3K, Akt protein and phosphorylated IRS2, PI3K, Akt protein were evaluated by Western Blot. All the data were analyzed by SPSS 17.0. Four weeks of treatment with LWDHT could significantly decrease the level of FBG and FINS in serum, improve the cellular morphology of liver, kidney, pancreas tissue, and the expression of IRS2, PI3K, Akt mRNA and phosphorylated IRS2, PI3K, Akt

  4. Sequential activation of phosphatidylinositol 3-kinase, beta Pix, Rac1, and Nox1 in growth factor-induced production of H2O2.

    PubMed

    Park, Hye Sun; Lee, Seung Hye; Park, Dongeun; Lee, Jun Sung; Ryu, Sung Ho; Lee, Won Jae; Rhee, Sue Goo; Bae, Yun Soo

    2004-05-01

    The generation of reactive oxygen species (ROS) in cells stimulated with growth factors requires the activation of phosphatidylinositol 3-kinase (PI3K) and the Rac protein. We report here that the COOH-terminal region of Nox1, a protein related to gp91(phox) (Nox2) of phagocytic cells, is constitutively associated with beta Pix, a guanine nucleotide exchange factor for Rac. Both growth factor-induced ROS production and Rac1 activation were completely blocked in cells depleted of beta Pix by RNA interference. Rac1 was also shown to bind to the COOH-terminal region of Nox1 in a growth factor-dependent manner. Moreover, the depletion of Nox1 by RNA interference inhibited growth factor-induced ROS generation. These results suggest that ROS production in growth factor-stimulated cells is mediated by the sequential activation of PI3K, beta Pix, and Rac1, which then binds to Nox1 to stimulate its NADPH oxidase activity.

  5. WAVE2, N-WASP, and Mena facilitate cell invasion via phosphatidylinositol 3-kinase-dependent local accumulation of actin filaments.

    PubMed

    Takahashi, Kazuhide; Suzuki, Katsuo

    2011-11-01

    Cell migration is accomplished by the formation of cellular protrusions such as lamellipodia and filopodia. These protrusions result from actin filament (F-actin) rearrangement at the cell cortex by WASP/WAVE family proteins and Drosophila enabled (Ena)/vasodilator-stimulated factor proteins. However, the role of each of these actin cytoskeletal regulatory proteins in the regulation of three-dimensional cell invasion remains to be clarified. We found that platelet-derived growth factor (PDGF) induces invasion of MDA-MB-231 human breast cancer cells through invasion chamber membrane pores. This invasion was accompanied by intensive F-actin accumulation at the sites of cell infiltration. After PDGF stimulation, WAVE2, N-WASP, and a mammalian Ena (Mena) colocalized with F-actin at the sites of cell infiltration in a phosphatidylinositol 3-kinase (PI3K)-dependent manner. Depletion of WAVE2, N-WASP, or Mena by RNA interference (RNAi) abrogated both cell invasion and intensive F-actin accumulation at the invasion site. These results indicate that by mediating intensive F-actin accumulation at the sites of cell infiltration, WAVE2, N-WASP, and Mena are crucial for PI3K-dependent cell invasion induced by PDGF.

  6. Mitogen-activated protein kinases and phosphatidylinositol 3-kinase are involved in Prevotella intermedia-induced proinflammatory cytokines expression in human periodontal ligament cells.

    PubMed

    Guan, Su-Min; Zhang, Ming; He, Jian-Jun; Wu, Jun-Zheng

    2009-08-28

    Chronic periodontitis is an inflammatory disease affecting periodontal connective tissues and alveolar bone. Proinflammatory mediators induced by periodontal pathogens play vital roles in the initiation and progression of the disease. In this study, we examined whether Prevotella intermedia induces proinflammatory cytokines expression in human periodontal ligament cells (hPDLs). The mRNA expression and protein production were determined by reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbant assay (ELISA) respectively. P. intermedia treatment dose- and time-dependently increased IL-6, IL-8 and M-CSF, but not IL-1beta and TNF-alpha mRNA expression and protein secretion. Preincubation of hPDLs with extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 kinase and phosphatidylinositol 3-kinase (PI3K) inhibitors PD98059, SP600125, SB203580 and LY294002 resulted in significant reduction in P. intermedia-induced IL-6, IL-8 and M-CSF expression. Blocking the synthesis of prostaglandin E(2) (PGE(2)) by indomethacin also abolished the stimulatory effects of P. intermedia on cytokines expression. Our results indicate that P. intermedia induces proinflammatory cytokines through MAPKs and PI3K signaling pathways, and PGE(2) is involved in the P. intermedia-induced proinflammatory cytokines upregulation.

  7. Neurotoxicity of developmental hypothyroxinemia and hypothyroidism in rats: Impairments of long-term potentiation are mediated by phosphatidylinositol 3-kinase signaling pathway.

    PubMed

    Wang, Yi; Wei, Wei; Wang, Yuan; Dong, Jing; Song, Binbin; Min, Hui; Teng, Weiping; Chen, Jie

    2013-09-01

    Neurotoxicity of iodine deficiency-induced hypothyroidism during developmental period results in serious impairments of brain function, such as learning and memory. These impairments are largely irreversible, and the underlying mechanisms remain unclear. In addition to hypothyroidism, iodine deficiency may cause hypothyroxinemia, a relatively subtle form of thyroid hormone deficiency. Neurotoxicity of developmental hypothyroxinemia also potentially impairs learning and memory. However, more direct evidence of the associations between developmental hypothyroxinemia and impairments of learning and memory should be provided, and the underlying mechanisms remain to be elucidated. Thus, in the present study, we investigated the effects of developmental hypothyroxinemia and hypothyroidism on long-term potentiation (LTP), a widely accepted cellular model of learning and memory, in the hippocampal CA1 region. The activation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway - a pathway closely associated with synaptic plasticity and learning and memory - was also investigated. Wistar rats were treated with iodine deficient diet or methimazole (MMZ) to induce developmental hypothyroxinemia or hypothyroidism. The results showed that developmental hypothyroxinemia caused by mild iodine deficiency and developmental hypothyroidism caused by severe iodine deficiency or MMZ significantly reduced the field-excitatory postsynaptic potential (f-EPSP) slope and the population spike (PS) amplitude. Decreased activation of the PI3K signaling pathway was also observed in rats subjected to developmental hypothyroxinemia or hypothyroidism. Our results may support the hypothesis that neurotoxicity of both developmental hypothyroxinemia and hypothyroidism causes damages to learning and memory. Our results also suggest that decreased activation of the PI3K signaling pathway may contribute to impairments of LTP caused by neurotoxicity of both developmental hypothyroxinemia and

  8. Effect of L-arginine supplementation on the hepatic phosphatidylinositol 3-kinase signaling pathway and gluconeogenic enzymes in early intrauterine growth-restricted rats.

    PubMed

    Luo, Kaiju; Chen, Pingyang; Li, Suping; Li, Wen; He, Mingfeng; Wang, Tao; Chen, Juncao

    2017-09-01

    The present study aimed to investigate the response of the phosphatidylinositol 3-kinase (PI3K) signaling pathway and gluconeogenic enzymes in intrauterine growth-restricted rats to dietary L-arginine (L-Arg) supplementation during the lactation period early in life. Pregnant Sprague-Dawley rats were randomly divided into a control group (CON), an intrauterine growth restriction group (IUGR) and an L-Arg group (LA). The pregnant rats in the CON group were fed a 21% protein diet, and those in the IUGR and LA groups were fed a 10% low protein diet, and all rats were fed a 21% protein diet after delivery. Water was available ad libitum to the pregnant rats during the 21-day lactation period, and the water provided to the LA group included 200 mg/kg/day L-Arg. Blood glucose, serum insulin, homeostasis model of assessment for insulin resistance (HOMA-IR), PI3K and protein kinase B (PKB) protein expression, and phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G-6-Pase) mRNA expression in the offspring rats were measured postnatally at 1, 3 and 8 weeks. No significant difference in blood glucose, serum insulin and HOMA-IR were identified at any time point among the three groups. PI3K and PKB expression was lower in the IUGR group offspring compared with that in the CON group offspring, but both were increased by dietary L-Arg supplementation. PEPCK mRNA and G-6-Pase mRNA expression levels in the offspring of the IUGR group were higher compared with those in the CON group but were downregulated following L-Arg supplementation. These results suggest that dietary L-Arg supplementation during the early lactation period promoted catch-up growth and reversed abnormalities in hepatic insulin signaling and gene expression of gluconeogenic enzymes in IUGR offspring rats.

  9. Inhibition of focal adhesion kinase induces apoptosis in bladder cancer cells via Src and the phosphatidylinositol 3-kinase/Akt pathway

    PubMed Central

    KONG, DEBO; CHEN, FENG; SIMA, NI

    2015-01-01

    Focal adhesion kinase (FAK) is a 125-kDa, cytosolic, non-receptor, protein tyrosine kinase localized at focal adhesions that can be activated by multiple inputs and in different manners. FAK is implicated in signaling pathways regulating cell movement, invasion, survival, gene expression and cancer stem cell self-renewal. The aim of the present study was to investigate whether FAK plays a role in the apoptosis of bladder cancer cells. The study employed in situ deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling and Annexin V labeling flow cytometry. It was found that both the knockdown of FAK and the suppression of FAK phosphorylation were able to induce apoptosis in bladder cancer cells. Caspase-3 was activated during the apoptosis induced by the suppression of FAK phosphorylation. Src was involved in FAK-regulated apoptosis in bladder cancer cells, while the suppression of Src phosphorylation was able to inhibit FAK tyrosine phosphorylation and induce apoptosis. Furthermore, phosphatidylinositol 3-kinase (PI3K)/Akt signaling was inhibited via the suppression of FAK tyrosine phosphorylation. Conversely, the expression of neither the general nor the tyrosine-phosphorylated FAK was regulated by inhibiting PI3K/Akt, which suggested that PI3K/Akt acted downstream of FAK to regulate apoptosis in bladder cancer cells. These findings indicate the presence of a mechanism of apoptosis involving FAK-mediated oncogenic signaling. FAK may function as an important regulator of extracellular signaling-mediated apoptosis in bladder cancer and be used as a novel therapeutic target in the treatment of the condition. PMID:26640543

  10. Simultaneous inhibition of mitogen-activated protein kinase and phosphatidylinositol 3-kinase pathways augment the sensitivity to actinomycin D in Ewing sarcoma.

    PubMed

    Yamamoto, Takatoshi; Ohno, Takatoshi; Wakahara, Kazuhiko; Nagano, Akihito; Kawai, Gou; Saitou, Mitsuru; Takigami, Iori; Matsuhashi, Aya; Yamada, Kazunari; Shimizu, Katsuji

    2009-08-01

    Ewing sarcoma cells, of which over 85% retain chimeric fusion gene EWS/Fli-1, are by and large more resistant to chemotherapeutics compared to nonneoplastic cells. The purpose of this study is to determine the role of EWS/Fli-1 fusion and its downstream targets regarding the cells' resistance against actinomycin D (ActD), which is one of the most commonly used antitumor agents in combination chemotherapy of Ewing sarcomas. Cytotoxicity was measured by WST-8 assay. Caspase-dependent and -independent cell death was examined by fluorescence microscope. Protein expression was analyzed by western blotting. Caspase activity was determined by Caspase-Glo assay. ActD-induced caspase-dependent apoptotic cell death to Ewing sarcoma TC-135 cells in a dose- and time- dependent manner. Knockdown of EWS/Fli-1 fusion by siRNA resulted in enhancement of ActD-induced apoptosis. ActD treatment activated both mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3-kinase (PI3K)/Akt pathways although in a distinctive manner. Combined administration of U0126 (MEK inhibitor) and LY294002 (PI3K inhibitor) significantly enhanced ActD-induced apoptosis in vitro and suppressed xenograft tumor growth in vivo. The present study demonstrated for the first time that combination of U0126 and LY294002 can augment the cytotoxicity of ActD against Ewing sarcoma cells in vitro and in vivo. Our results indicate that further study on combination of conventional chemotherapies with MEK and PI3K inhibitors may be considered for innovative treatments of Ewing sarcoma patients.

  11. Curcumin modulates nuclear factor kappaB (NF-kappaB)-mediated inflammation in human tenocytes in vitro: role of the phosphatidylinositol 3-kinase/Akt pathway.

    PubMed

    Buhrmann, Constanze; Mobasheri, Ali; Busch, Franziska; Aldinger, Constance; Stahlmann, Ralf; Montaseri, Azadeh; Shakibaei, Mehdi

    2011-08-12

    Inflammatory processes play essential roles in the pathogenesis of tendinitis and tendinopathy. These events are accompanied by catabolic processes initiated by pro-inflammatory cytokines such as interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). Pharmacological treatments for tendinitis are restricted to the use of non-steroidal anti-inflammatory drugs. Recent studies in various cell models have demonstrated that curcumin targets the NF-κB signaling pathway. However, its potential for the treatment of tendinitis has not been explored. Herein, we used an in vitro model of human tenocytes to study the mechanism of curcumin action on IL-1β-mediated inflammatory signaling. Curcumin at concentrations of 5-20 μm inhibited IL-1β-induced inflammation and apoptosis in cultures of human tenocytes. The anti-inflammatory effects of curcumin included down-regulation of gene products that mediate matrix degradation (matrix metalloproteinase-1, -9, and -13), prostanoid production (cyclooxygenase-2), apoptosis (Bax and activated caspase-3), and stimulation of cell survival (Bcl-2), all known to be regulated by NF-κB. Furthermore, curcumin suppressed IL-1β-induced NF-κB activation via inhibition of phosphorylation and degradation of inhibitor of κBα, inhibition of inhibitor of κB-kinase activity, and inhibition of nuclear translocation of NF-κB. Furthermore, the effects of IL-1β were abrogated by wortmannin, suggesting a role for the phosphatidylinositol 3-kinase (PI-3K) pathway in IL-1β signaling. Curcumin suppressed IL-1β-induced PI-3K p85/Akt activation and its association with IKK. These results demonstrate, for the first time, a potential role for curcumin in treating tendon inflammation through modulation of NF-κB signaling, which involves PI-3K/Akt and the tendon-specific transcription factor scleraxis in tenocytes.

  12. Safety and Efficacy of Buparlisib (BKM120) in Patients with PI3K Pathway-Activated Non-Small Cell Lung Cancer: Results from the Phase II BASALT-1 Study.

    PubMed

    Vansteenkiste, Johan F; Canon, Jean-Luc; De Braud, Filippo; Grossi, Francesco; De Pas, Tommaso; Gray, Jhanelle E; Su, Wu-Chou; Felip, Enriqueta; Yoshioka, Hiroshige; Gridelli, Cesare; Dy, Grace K; Thongprasert, Sumitra; Reck, Martin; Aimone, Paola; Vidam, Gena Atalla; Roussou, Pantelia; Wang, Ying A; Di Tomaso, Emmanuelle; Soria, Jean-Charles

    2015-09-01

    The phosphatidylinositol 3-kinase (PI3K) pathway promotes tumor growth and treatment resistance in non-small cell lung cancer (NSCLC). The aim of the open-label, two-stage, Phase II study BASALT-1 (NCT01820325) was to investigate the pan-PI3K inhibitor buparlisib (BKM120) in patients with PI3K pathway-activated, relapsed NSCLC. After prescreening for PI3K pathway activation, patients with PI3K pathway-activated, metastatic, squamous or nonsquamous NSCLC, who had relapsed after prior systemic antineoplastic therapy, were enrolled. In Stage 1, patients received single-agent buparlisib (100 mg/day). A futility analysis was performed independently in each histology group, based on the 12-week progression-free survival rate for the first 30 patients treated in each group being less than 50%. Exploratory biomarker analyses were performed in archival tissue samples and circulating tumor DNA (ctDNA). Of 1242 prescreened patients, 13.5% exhibited PI3K pathway activation. As of June 5, 2014, 63 patients (30 squamous and 33 nonsquamous) were treated in Stage 1. The 12-week progression-free survival rates were 23.3% (95% confidence interval: 9.9-42.3) and 20.0% (95% confidence interval: 7.7-38.6) in the squamous and nonsquamous groups, respectively. Stage 2 was therefore not initiated in either group. PI3K pathway mutations in ctDNA were more concordant with metastatic tissue than with primary biopsies. Despite preselecting patients for targeted treatment, BASALT-1 did not meet its primary objective during Stage 1. PI3K pathway activation can be detected using ctDNA, but may not be the main oncogenic driver in NSCLC. Combinations of PI3K inhibitors with other agents may demonstrate greater efficacy than monotherapy.

  13. GS-1101: a delta-specific PI3K inhibitor in chronic lymphocytic leukemia.

    PubMed

    Macias-Perez, Ines M; Flinn, Ian W

    2013-03-01

    Chronic lymphocytic leukemia (CLL) remains an incurable B-cell malignancy with many unanswered questions. While the cell of origin and etiology are still unknown, significant scientific progress has revealed numerous molecular targets for novel therapeutic interventions. Phosphatidylinositol 3-kinases (PI3K) regulate key cellular functions, including growth, survival and migration, by integrating and transmitting signals from diverse surface molecules including the B-cell receptor (BCR). In lymphocytes, the PI3Kδ isoform plays a critical role in B-cell homeostasis and function. In CLL, the PI3K pathway is constitutively active and dependent on PI3Kδ. GS-1101 is a highly selective PI3Kδ inhibitor that in CLL patients causes a rapid and sustained reduction in lymphadenopathy, accompanied by transient lymphocytosis. This article will review new insights into the pathophysiology of CLL, the preclinical rationale of a PI3Kδ inhibitor in CLL, and the clinical evidence supporting this first-in-class therapeutic target for CLL patients.

  14. PI3K: An Attractive Candidate for the Central Integration of Metabolism and Reproduction

    PubMed Central

    Acosta-Martínez, Maricedes

    2012-01-01

    In neurons, as in a variety of other cell types, the enzyme phosphatidylinositol-3-kinase (PI3K) is a key intermediate that is common to the signaling pathways of a number of peripheral metabolic cues, including insulin and leptin, which are well known to regulate both metabolic and reproductive functions. This review article will explore the possibility that PI3K is a key integrator of metabolic and neural signals regulating gonadotropin releasing hormone (GnRH)/luteinizing hormone (LH) release and explore the hypothesis that this enzyme is pivotal in many disorders where gonadotropin release is at risk. Although the mechanisms mediating the influence of metabolism and nutrition on fertility are currently unclear, the strong association between metabolic disorders and infertility is undeniable. For example, women suffering from anorectic disorders experience amenorrhea as a consequence of malnutrition-induced impairment of LH release, and at the other extreme, obesity is also commonly co-morbid with menstrual dysfunction and infertility. Impaired hypothalamic insulin and leptin receptor signaling is thought to be at the core of reproductive disorders associated with metabolic dysfunction. While low levels of leptin and insulin characterize states of negative energy balance, prolonged nutrient excess is associated with insulin and leptin resistance. Metabolic models known to alter GnRH/LH release such as diabetes, diet-induced obesity, and caloric restriction are also accompanied by impairment of PI3K signaling in insulin and leptin sensitive tissues including the hypothalamus. However, a clear link between this signaling pathway and the control of GnRH release by peripheral metabolic cues has not been established. Investigating the role of the signaling pathways shared by metabolic cues that are critical for a normal reproductive state can help identify possible targets in the treatment of metabolic and reproductive disorders such as polycystic ovarian syndrome

  15. PI3K: An Attractive Candidate for the Central Integration of Metabolism and Reproduction.

    PubMed

    Acosta-Martínez, Maricedes

    2011-01-01

    In neurons, as in a variety of other cell types, the enzyme phosphatidylinositol-3-kinase (PI3K) is a key intermediate that is common to the signaling pathways of a number of peripheral metabolic cues, including insulin and leptin, which are well known to regulate both metabolic and reproductive functions. This review article will explore the possibility that PI3K is a key integrator of metabolic and neural signals regulating gonadotropin releasing hormone (GnRH)/luteinizing hormone (LH) release and explore the hypothesis that this enzyme is pivotal in many disorders where gonadotropin release is at risk. Although the mechanisms mediating the influence of metabolism and nutrition on fertility are currently unclear, the strong association between metabolic disorders and infertility is undeniable. For example, women suffering from anorectic disorders experience amenorrhea as a consequence of malnutrition-induced impairment of LH release, and at the other extreme, obesity is also commonly co-morbid with menstrual dysfunction and infertility. Impaired hypothalamic insulin and leptin receptor signaling is thought to be at the core of reproductive disorders associated with metabolic dysfunction. While low levels of leptin and insulin characterize states of negative energy balance, prolonged nutrient excess is associated with insulin and leptin resistance. Metabolic models known to alter GnRH/LH release such as diabetes, diet-induced obesity, and caloric restriction are also accompanied by impairment of PI3K signaling in insulin and leptin sensitive tissues including the hypothalamus. However, a clear link between this signaling pathway and the control of GnRH release by peripheral metabolic cues has not been established. Investigating the role of the signaling pathways shared by metabolic cues that are critical for a normal reproductive state can help identify possible targets in the treatment of metabolic and reproductive disorders such as polycystic ovarian syndrome.

  16. Different inhibition of Gβγ-stimulated class IB phosphoinositide 3-kinase (PI3K) variants by a monoclonal antibody. Specific function of p101 as a Gβγ-dependent regulator of PI3Kγ enzymatic activity.

    PubMed

    Shymanets, Aliaksei; Prajwal; Vadas, Oscar; Czupalla, Cornelia; LoPiccolo, Jaclyn; Brenowitz, Michael; Ghigo, Alessandra; Hirsch, Emilio; Krause, Eberhard; Wetzker, Reinhard; Williams, Roger L; Harteneck, Christian; Nürnberg, Bernd

    2015-07-01

    Class IB phosphoinositide 3-kinases γ (PI3Kγ) are second-messenger-generating enzymes downstream of signalling cascades triggered by G-protein-coupled receptors (GPCRs). PI3Kγ variants have one catalytic p110γ subunit that can form two different heterodimers by binding to one of a pair of non-catalytic subunits, p87 or p101. Growing experimental data argue for a different regulation of p87-p110γ and p101-p110γ allowing integration into distinct signalling pathways. Pharmacological tools enabling distinct modulation of the two variants are missing. The ability of an anti-p110γ monoclonal antibody [mAb(A)p110γ] to block PI3Kγ enzymatic activity attracted us to characterize this tool in detail using purified proteins. In order to get insight into the antibody-p110γ interface, hydrogen-deuterium exchange coupled to MS (HDX-MS) measurements were performed demonstrating binding of the monoclonal antibody to the C2 domain in p110γ, which was accompanied by conformational changes in the helical domain harbouring the Gβγ-binding site. We then studied the modulation of phospholipid vesicles association of PI3Kγ by the antibody. p87-p110γ showed a significantly reduced Gβγ-mediated phospholipid recruitment as compared with p101-p110γ. Concomitantly, in the presence of mAb(A)p110γ, Gβγ did not bind to p87-p110γ. These data correlated with the ability of the antibody to block Gβγ-stimulated lipid kinase activity of p87-p110γ 30-fold more potently than p101-p110γ. Our data argue for differential regulatory functions of the non-catalytic subunits and a specific Gβγ-dependent regulation of p101 in PI3Kγ activation. In this scenario, we consider the antibody as a valuable tool to dissect the distinct roles of the two PI3Kγ variants downstream of GPCRs.

  17. Activation of phosphatidylinositol 3-kinase/Akt-mammalian target of Rapamycin signaling pathway in the hippocampus is essential for the acquisition of morphine-induced place preference in rats.

    PubMed

    Cui, Yue; Zhang, X Q; Cui, Y; Xin, W J; Jing, J; Liu, X G

    2010-11-24

    Hippocampus is a critical structure for the acquisition of morphine-induced conditioned place preference (CPP), which is a usual learning paradigm for assessing drug reward. However, the precise mechanisms remain largely unknown. Phosphatidylinositol 3-kinase (PI3K) and its downstream targets, including Akt, mammalian target of Rapamycin (mTOR) and 70-kDa ribosomal S6 kinase (p70S6K), are critical molecules implicated in learning and memory. Here, we tested the role of PI3K/Akt-mTOR-p70S6K signaling pathway in morphine-induced CPP in the hippocampus. Our results showed that the acquisition of morphine CPP increased phosphorylation of Akt in the hippocampal CA3, but not in the nucleus accumbens (NAc), the ventral tegmental area (VTA) or the CA1. Moreover, the phosphorylated Akt exclusively expressed in the CA3 neurons. Likewise, levels of phosphorylated mTOR and p70S6K were significantly enhanced in the CA3 following morphine CPP. The alterations of these phosphorylated proteins are positively correlated with the acquisition of morphine CPP. More importantly, microinjection of PI3K inhibitor (LY294002) or mTOR inhibitor (Rapamycin) into the CA3 prevented the acquisition of CPP and inhibited the activation of PI3K-Akt signaling pathway. In addition, pre-infusion of β-FNA (β-funaltrexamine hydrochloride), a selective irreversible μ opioid receptor antagonist, into CA3 significantly prevented the acquisition of CPP and impaired Akt phosphorylation. All these results strongly implied that the PI3K-Akt signaling pathway activated by μ opioid receptor in hippocampal CA3 plays an important role in acquisition of morphine-induced CPP.

  18. Roles of phosphatidylinositol 3-kinase and p38 mitogen-activated protein kinase in the regulation of protein kinase C-alpha activation in interferon-gamma-stimulated macrophages.

    PubMed

    Hardy, Pierre-Olivier; Diallo, Tamsir O; Matte, Christine; Descoteaux, Albert

    2009-09-01

    Members of the protein kinase C (PKC) family are activated by interferon-gamma (IFN-gamma) and modulate IFN-gamma-induced cellular responses by regulating the activity of transcription factors. We previously reported that PKC-alpha enhances the ability of IFN regulatory factor-1 to transactivate the class II transactivator (CIITA) promoter IV in IFN-gamma-stimulated macrophages. In addition, we showed that IFN-gamma induces the nuclear translocation of PKC-alpha but the mechanisms for this remain to be elucidated. In this study, we sought to identify signalling pathways involved in IFN-gamma-induced activation of PKC-alpha and to characterize their potential roles in modulating IFN-gamma-induced responses in macrophages. IFN-gamma-mediated nuclear translocation of PKC-alpha was a Janus activated kinase 2 (JAK2)-independent process, which required phosphatidylinositol 3-kinase (PI3K) and p38 mitogen-activated protein kinase (MAPK). However, PKC-alpha phosphorylation was independent of PI3K and p38 MAPK, indicating that IFN-gamma-induced phosphorylation and nuclear translocation of PKC-alpha are mediated by distinct mechanisms. In addition, inhibition of PI3K, but not of p38 MAPK, strongly impaired IFN-gamma-induced CIITA and MHC II gene expression. Finally, PKC-alpha associated with signal transducer and activator of transcription 1 (STAT1) and was required for the phosphorylation of STAT1 on serine 727 in IFN-gamma-stimulated macrophages. Taken together, our data indicate that PI3K and p38 MAPK modulate IFN-gamma-stimulated PKC-alpha nuclear translocation independently of JAK2 activity and that both PI3K and PKC-alpha are required for type IV CIITA and MHC II gene expression in IFN-gamma-stimulated macrophages.

  19. Phosphatidylinositol 3-kinase, phosphoinositide-specific phospholipase-Cgamma and protein kinase-C signal myelin phagocytosis mediated by complement receptor-3 alone and combined with scavenger receptor-AI/II in macrophages.

    PubMed

    Makranz, Chen; Cohen, Goni; Baron, Ayellet; Levidor, Lital; Kodama, Tatsuhiko; Reichert, Fanny; Rotshenker, Shlomo

    2004-03-01

    Complement-receptor-3 (CR3/MAC-1), scavenger-receptor-AI/II (SRAI/II) and Fcgamma-receptor (FcgammaR) can mediate phagocytosis of degenerated myelin in macrophages and microglia. However, CR3/MAC-1 and SRAI/II, but not FcgammaR, mediate phagocytosis after axonal injury. We tested for phosphatidylinositol 3-kinase (PI3K), phosphoinositide-specific phospholipase-Cgamma (PLCgamma) and protein kinase-C (PKC) signaling in myelin phagocytosis mediated by CR3/MAC-1 alone and by CR3/MAC-1 combined with SRAI/II. Phagocytosis was inhibited by PI3K inhibitors wortmannin and LY-294002, PLCgamma inhibitor U-73122, classical PKC (cPKC) inhibitor Go-6976, general PKC inhibitors Ro-318220 and calphostin-C, and BAPTA/AM which chelates intracellular Ca(2+) required for cPKC activation. PKC activator PMA augmented phagocytosis and further alleviated inhibitions induced by PI3K and PLCgamma inhibitors. Overall, altering PKC activity modulated phagocytosis 4- to 6-fold between inhibition and augmentation. PLCgamma activation did not require tyrosine phosphorylation. Thus, signaling of myelin phagocytosis mediated by CR3/MAC-1 alone and by CR3/MAC-1 combined with SRAI/II involves PI3K, PLCgamma and cPKC, the cascade PI3K-->PLCgamma-->cPKC, and wide-range modulation by PKC. This pathway may thus be targeted for in vivo modulation, which may explain differences in the efficiency of CR3/MAC-1-mediated myelin phagocytosis in different pathological conditions.

  20. Triptolide, a diterpenoid triepoxide, induces antitumor proliferation via activation of c-Jun NH{sub 2}-terminal kinase 1 by decreasing phosphatidylinositol 3-kinase activity in human tumor cells

    SciTech Connect

    Miyata, Yoshiki; Sato, Takashi . E-mail: satotak@ps.toyaku.ac.jp; Ito, Akira

    2005-11-04

    Triptolide, a diterpenoid triepoxide extracted from the Chinese herb Tripterygium wilfordii Hook f., exerts antitumorigenic actions against several tumor cells, but the intracellular target signal molecule(s) for this antitumorigenesis activity of triptolide remains to be identified. In the present study, we demonstrated that triptolide, in a dose-dependent manner, inhibited the proliferation of human fibrosarcoma HT-1080, human squamous carcinoma SAS, and human uterine cervical carcinoma SKG-II cells. In addition, triptolide was found to decrease phosphatidylinositol 3-kinase (PI3K) activity. A PI3K inhibitor, LY-294002, mimicked the triptolide-induced antiproliferative activity in HT-1080, SAS, and SKG-II cells. There was no change in the activity of Akt or protein kinase C (PKC), both of which are downstream effectors in the PI3K pathway. Furthermore, the phosphorylation of Ras, Raf, and mitogen-activated protein/extracellular signal-regulated kinase 1/2 was not modified in HT-1080 cells treated with triptolide. However, the phosphorylation of c-Jun NH{sub 2}-terminal kinase 1 (JNK1) was found to increase in both triptolide- and LY-294002-treated cells. Furthermore, the triptolide-induced inhibition of HT-1080 cell proliferation was not observed by JNK1 siRNA-treatment. These results provide novel evidence that PI3K is a crucial target molecule in the antitumorigenic action of triptolide. They further suggest a possible triptolide-induced inhibitory signal for tumor cell proliferation that is initiated by the decrease in PI3K activity, which in turn leads to the augmentation of JNK1 phosphorylation via the Akt and/or PKC-independent pathway(s). Moreover, it is likely that the activation of JNK1 is required for the triptolide-induced inhibition of tumor proliferation.

  1. PI3K delta and PI3K gamma: partners in crime in inflammation in rheumatoid arthritis and beyond?

    PubMed

    Rommel, Christian; Camps, Montserrat; Ji, Hong

    2007-03-01

    Dysregulated signal transduction in innate and adaptive immune cells is known to be associated with the development of various autoimmune and inflammatory diseases. Consequently, targeting intracellular signalling of the pro-inflammatory cytokine network heralds hope for the next generation of anti-inflammatory drugs. Phosphoinositide 3-kinases (PI3Ks) generate lipid-based second messengers that control an array of intracellular signalling pathways that are known to have important roles in leukocytes. In light of the recent progress in the development of selective PI3K inhibitors, and the beneficial effects of these inhibitors in models of acute and chronic inflammatory disorders, we discuss the therapeutic potential of blocking PI3K isoforms for the treatment of rheumatoid arthritis and other immune-mediated diseases.

  2. ARID1A Mutations and PI3K/AKT Pathway Alterations in Endometriosis and Endometriosis-Associated Ovarian Carcinomas

    PubMed Central

    Samartzis, Eleftherios P.; Noske, Aurelia; Dedes, Konstantin J.; Fink, Daniel; Imesch, Patrick

    2013-01-01

    Endometriosis is a common gynecological disease affecting 6%–10% of women of reproductive age and is characterized by the presence of endometrial-like tissue in localizations outside of the uterine cavity as, e.g., endometriotic ovarian cysts. Mainly, two epithelial ovarian carcinoma subtypes, the ovarian clear cell carcinomas (OCCC) and the endometrioid ovarian carcinomas (EnOC), have been molecularly and epidemiologically linked to endometriosis. Mutations in the gene encoding the AT-rich interacting domain containing protein 1A (ARID1A) have been found to occur in high frequency in OCCC and EnOC. The majority of these mutations lead to a loss of expression of the ARID1A protein, which is a subunit of the SWI/SNF chromatin remodeling complex and considered as a bona fide tumor suppressor. ARID1A mutations frequently co-occur with mutations, leading to an activation of the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, such as mutations in PIK3CA encoding the catalytic subunit, p110α, of PI3K. In combination with recent functional observations, these findings strongly suggest cooperating mechanisms between the two pathways. The occurrence of ARID1A mutations and alterations in the PI3K/AKT pathway in endometriosis and endometriosis-associated ovarian carcinomas, as well as the possible functional and clinical implications are discussed in this review. PMID:24036443

  3. Involvement of PI3K and MAPK Signaling in bcl-2-induced Vascular Endothelial Growth Factor Expression in Melanoma Cells

    PubMed Central

    Trisciuoglio, Daniela; Iervolino, Angela; Zupi, Gabriella; Del Bufalo, Donatella

    2005-01-01

    We have previously demonstrated that bcl-2 overexpression in tumor cells exposed to hypoxia increases the expression of vascular endothelial growth factor (VEGF) gene through the hypoxia-inducible factor-1 (HIF-1). In this article, we demonstrate that exposure of bcl-2 overexpressing melanoma cells to hypoxia induced phosphorylation of AKT and extracellular signal-regulated kinase (ERK)1/2 proteins. On the contrary, no modulation of these pathways by bcl-2 was observed under normoxic conditions. When HIF-1α expression was reduced by RNA interference, AKT and ERK1/2 phosphorylation were still induced by bcl-2. Pharmacological inhibition of mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) signaling pathways reduced the induction of VEGF and HIF-1 in response to bcl-2 overexpression in hypoxia. No differences were observed between control and bcl-2-overexpressing cells in normoxia, in terms of VEGF protein secretion and in response to PI3K and MAPK inhibitors. We also demonstrated that RNA interference-mediated down-regulation of bcl-2 expression resulted in a decrease in the ERK1/2 phosphorylation and VEGF secretion only in bcl-2-overexpressing cell exposed to hypoxia but not in control cells. In conclusion, our results indicate, for the first time, that bcl-2 synergizes with hypoxia to promote expression of angiogenesis factors in melanoma cells through both PI3K- and MAPK-dependent pathways. PMID:15987743

  4. CD38 is highly expressed and affects the PI3K/Akt signaling pathway in cervical cancer.

    PubMed

    Liao, Shan; Xiao, Songshu; Zhu, Guangchao; Zheng, Danwei; He, Junyu; Pei, Zhen; Li, Guiyuan; Zhou, Yanhong

    2014-12-01

    Cervical cancer is the second most common cancer and the fifth most deadly malignancy in females worldwide, affecting 500,000 individuals each year. It is the leading cause of cancer mortality among women in developing countries. Dysregulated activation of genes, such as CD44, SOX9 and SKP2, plays a role in cervical cancer. CD38 is known to be involved in activities typical of cell surface receptors, such as signaling for activation and proliferation events and heterotypic cell adhesion. CD38 contributes to disease progression and relapse in certain tumors, such as acute myeloid and chronic lymphocytic leukemia. To the best of our knowledge, there is currently no report on the relationship between CD38 and cervical cancer. Using qPCR, immunohistochemistry, and western blot analysis, the expression levels of CD38 were investigated and found to be upregulated in cervical cancer. CD38 was correlated with dysregulation of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway in cervical cancer tissues in vitro. At the same time, CD38 overexpression affected the expression of PI3K, Akt, MDM2 and p53 in vivo. The results of the present study suggested that CD38 is highly expressed in cervical carcinoma tissues and play an important role in dysregulation of the PI3K/Akt signaling pathway.

  5. Transforming growth factor alpha promotes osteosarcoma metastasis by ICAM-1 and PI3K/Akt signaling pathway.

    PubMed

    Hou, Chun-Han; Lin, Feng-Ling; Tong, Kai-Biao; Hou, Sheng-Mon; Liu, Ju-Fang

    2014-06-15

    Osteosarcoma is the most common primary malignancy of bone and is characterized by a high malignant and metastatic potential. Transforming growth factor alpha (TGF-α) is classified as the EGF (epidermal growth factor)-like family, which is involved in cancer cellular activities such as proliferation, motility, migration, adhesion and invasion abilities. However, the effect of TGF-α on human osteosarcoma is largely unknown. We found that TGF-α increased the cell migration and expression of intercellular adhesion molecule-1 (ICAM-1) in human osteosarcoma cells. Transfection of cells with ICAM-1 siRNA reduced TGF-α-mediated cell migration. We also found that the phosphatidylinositol 3'-kinase (PI3K)/Akt/NF-κB pathway was activated after TGF-α treatment, and TGF-α-induced expression of ICAM-1 and cell migration was inhibited by the specific inhibitors and siRNAs of PI3K, Akt, and NF-κB cascades. In addition, knockdown of TGF-α expression markedly decreased cell metastasis in vitro and in vivo. Our results indicate that TGF-α/EGFR interaction elicits PI3K and Akt activation, which in turn activates NF-κB, resulting in the expression of ICAM-1 and contributing the migration of human osteosarcoma cells.

  6. Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes.

    PubMed

    Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

    2014-11-01

    In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes.

  7. Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes

    PubMed Central

    Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

    2014-01-01

    In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes. PMID:25489416

  8. Consensus model for identification of novel PI3K inhibitors in large chemical library

    NASA Astrophysics Data System (ADS)

    Liew, Chin Yee; Ma, Xiao Hua; Yap, Chun Wei

    2010-02-01

    Phosphoinositide 3-kinases (PI3Ks) inhibitors have treatment potential for cancer, diabetes, cardiovascular disease, chronic inflammation and asthma. A consensus model consisting of three base classifiers (AODE, kNN, and SVM) trained with 1,283 positive compounds (PI3K inhibitors), 16 negative compounds (PI3K non-inhibitors) and 64,078 generated putative negatives was developed for predicting compounds with PI3K inhibitory activity of IC50 ≤ 10 μM. The consensus model has an estimated false positive rate of 0.75%. Nine novel potential inhibitors were identified using the consensus model and several of these contain structural features that are consistent with those found to be important for PI3K inhibitory activities. An advantage of the current model is that it does not require knowledge of 3D structural information of the various PI3K isoforms, which is not readily available for all isoforms.

  9. Differential Phosphatidylinositol-3-Kinase-Akt-mTOR Activation by Semliki Forest and Chikungunya Viruses Is Dependent on nsP3 and Connected to Replication Complex Internalization

    PubMed Central

    Biasiotto, Roberta; Eng, Kai; Neuvonen, Maarit; Götte, Benjamin; Rheinemann, Lara; Mutso, Margit; Utt, Age; Varghese, Finny; Balistreri, Giuseppe; Merits, Andres; Ahola, Tero; McInerney, Gerald M.

    2015-01-01

    ABSTRACT Many viruses affect or exploit the phosphatidylinositol-3-kinase (PI3K)-Akt-mammalian target of rapamycin (mTOR) pathway, a crucial prosurvival signaling cascade. We report that this pathway was strongly activated in cells upon infection with the Old World alphavirus Semliki Forest virus (SFV), even under conditions of complete nutrient starvation. We mapped this activation to the hyperphosphorylated/acidic domain in the C-terminal tail of SFV nonstructural protein nsP3. Viruses with a deletion of this domain (SFV-Δ50) but not of other regions in nsP3 displayed a clearly delayed and reduced capacity of Akt stimulation. Ectopic expression of the nsP3 of SFV wild type (nsP3-wt), but not nsP3-Δ50, equipped with a membrane anchor was sufficient to activate Akt. We linked PI3K-Akt-mTOR stimulation to the intracellular dynamics of viral replication complexes, which are formed at the plasma membrane and subsequently internalized in a process blocked by the PI3K inhibitor wortmannin. Replication complex internalization was observed upon infection of cells with SFV-wt and SFV mutants with deletions in nsP3 but not with SFV-Δ50, where replication complexes were typically accumulated at the cell periphery. In cells infected with the closely related chikungunya virus (CHIKV), the PI3K-Akt-mTOR pathway was only moderately activated. Replication complexes of CHIKV were predominantly located at the cell periphery. Exchanging the hypervariable C-terminal tail of nsP3 between SFV and CHIKV induced the phenotype of strong PI3K-Akt-mTOR activation and replication complex internalization in CHIKV. In conclusion, infection with SFV but not CHIKV boosts PI3K-Akt-mTOR through the hyperphosphorylated/acidic domain of nsP3 to drive replication complex internalization. IMPORTANCE SFV and CHIKV are very similar in terms of molecular and cell biology, e.g., regarding replication and molecular interactions, but are strikingly different regarding pathology: CHIKV is a relevant human

  10. Inhibition of Phosphatidylinositol 3-Kinase/Akt Signaling Suppresses Tumor Cell Proliferation and Neuroendocrine Marker Expression in GI Carcinoid Tumors

    PubMed Central

    Pitt, Susan C.; Chen, Herbert; Kunnimalaiyaan, Muthusamy

    2010-01-01

    Background Over-activation of PI3K/Akt signaling facilitates tumor proliferation in several cancers. We have shown that various signal transduction pathways promote tumorigenesis in carcinoid tumors, which exhibit endogenously high levels of active, phosphorylated Akt. Therefore, we hypothesized that inhibition of the PI3K/Akt pathway would suppress carcinoid tumor cell growth and neuroendocrine (NE) marker production. Methods Human carcinoid BON cells were treated in vitro with LY294002, a PI3 kinase inhibitor, or transfected with Akt1 siRNA. Tumor cell proliferation was measured by MTT for six days. The effect of LY294002 or Akt1 siRNA treatment was assessed by western analysis. We examined the levels of phosphorylated Akt, total Akt, Akt1, and the NE markers human achaete-scute homolog1 (ASCL1) and chromogranin A (CgA). Results Treatment of BON cells with LY294002 reduced tumor cell proliferation (76%) in a dose-dependent manner. Growth also decreased in Akt1 siRNA transfected cells (29%). Levels of active, phosphorylated Akt and the NE tumor markers, ASCL1 and CgA, were diminished with both LY294002 and Akt1 siRNA treatments proportional to the degree of Akt inhibition. Total Akt, Akt2, and Akt3 levels were unaffected by these experiments. Conclusions These data indicate that PI3K/Akt signaling performs a critical role in human carcinoid tumor cell survival and NE hormone generation. Furthermore, the development of novel therapeutics targeting Akt1 or components of the PI3K/Akt pathway may enhance the management of carcinoid disease. Synopsis Carcinoid tumor cells were treated with a PI3K inhibitor, LY294002, and Akt1 siRNA to delineate the role of PI3K/Akt signaling in carcinoids. The effects of treatment on cellular proliferation and neuroendocrine marker expression were observed. PMID:19588205

  11. Colon Cancer Tumorigenesis Initiated by the H1047R Mutant PI3K

    PubMed Central

    Yueh, Alexander E.; Payne, Susan N.; Leystra, Alyssa A.; Van De Hey, Dana R.; Foley, Tyler M.; Pasch, Cheri A.; Clipson, Linda; Matkowskyj, Kristina A.; Deming, Dustin A.

    2016-01-01

    The phosphoinositide 3-kinase (PI3K) signaling pathway is critical for multiple important cellular functions, and is one of the most commonly altered pathways in human cancers. We previously developed a mouse model in which colon cancers were initiated by a dominant active PI3K p110-p85 fusion protein. In that model, well-differentiated mucinous adenocarcinomas developed within the colon and initiated through a non-canonical mechanism that is not dependent on WNT signaling. To assess the potential relevance of PI3K mutations in human cancers, we sought to determine if one of the common mutations in the human disease could also initiate similar colon cancers. Mice were generated expressing the Pik3caH1047R mutation, the analog of one of three human hotspot mutations in this gene. Mice expressing a constitutively active PI3K, as a result of this mutation, develop invasive adenocarcinomas strikingly similar to invasive adenocarcinomas found in human colon cancers. These tumors form without a polypoid intermediary and also lack nuclear CTNNB1 (β-catenin), indicating a non-canonical mechanism of tumor initiation mediated by the PI3K pathway. These cancers are sensitive to dual PI3K/mTOR inhibition indicating dependence on the PI3K pathway. The tumor tissue remaining after treatment demonstrated reduction in cellular proliferation and inhibition of PI3K signaling. PMID:26863299

  12. Role of PI3-K/Akt pathway and its effect on glial cell line-derived neurotrophic factor in midbrain dopamine cells.

    PubMed

    Wang, Hong-jun; Cao, Jun-ping; Yu, Jing-kao; Gao, Dian-shuai

    2007-02-01

    To explore the intracellular mechanisms underlying the survival/differentiation effect of the glial cell line-derived neurotrophic factor (GDNF) on dopamine (DA) cells. Midbrain slice culture and primary cell culture were established, and the cultures were divided into 3 groups: control group, GDNF group, and the phosphatidylinositol 3-kinase/Akt (PI3-K/Akt) pathway-inhibited group. Then the expression of tyrosine hydroxylase (TH) was detected by immunostaining as well as Western blotting. GDNF treatment induced an increase in the number of TH-immunoreactive (ir) cells and the neurite number of TH-ir cells, as well as in the level of TH expression in cultures (Number of TH-ir cells in the slice culture: control group, 8.76+/-0.75; GDNF group, 18.63+/-0.95. Number of TH-ir cells and neurite number of TH-ir cells in cell culture: control group, 3.65+/-0.88 and 2.49+/-0.42; GDNF group, 6.01+/-0.43 and 4.89+/-0.46). Meanwhile, the stimulation of cultured cells with GDNF increased the phosphorylation of Akt, which is a downstream effector of PI3-K/Akt. The effects of GDNF were specifically blocked by the inhibitor of the PI3-K/Akt pathway, wortmannin (Number of TH-ir cells in slice culture: PI3-K/Akt pathway-inhibited group, 6.98+/-0.58. Number of TH-ir cells and neurite number of TH-ir cells in cell culture: PI3-K/Akt pathway-inhibited group, 3.79+/-0.62 and 2.50+/-0.25, respectively). The PI3-K/Akt pathway mediates the survival/differentiation effect of GDNF on DA cells.

  13. An essential role for the Id1/PI3K/Akt/NFkB/survivin signalling pathway in promoting the proliferation of endothelial progenitor cells in vitro.

    PubMed

    Li, Wei; Wang, Hang; Kuang, Chun-Yan; Zhu, Jin-Kun; Yu, Yang; Qin, Zhe-Xue; Liu, Jie; Huang, Lan

    2012-04-01

    The enhancement of re-endothelialisation is a critical therapeutic option for repairing injured blood vessels. Endothelial progenitor cells (EPCs) are the major source of cells that participate in endothelium repair and contribute to re-endothelialisation by reducing neointima formation after vascular injury. The over-expression of the inhibitor of differentiation or DNA binding 1 (Id1) significantly improved EPC proliferation. This study aimed to investigate the effects of Id1 on the phosphatidylinositol-3-kinase (PI3K)/Akt/nuclear factor kappa B (NFκB)/survivin signalling pathway and its significance in promoting EPC proliferation in vitro. Spleen-derived EPCs were cultured as previously described. Id1 was presented at low levels in EPCs, and was rapidly up-regulated by stimulation with vascular endothelial growth factor. We demonstrated that transient transfection of Id1 into EPCs activated the PI3K/Akt/NFκB/survivin signalling pathway and promoted EPC proliferation. The proliferation of EPCs was extensively inhibited by silencing of endogenous Id1, and knockdown of Id1 expression led to suppression of PI3K/Akt/NFκB/survivin signalling pathway in EPCs. In addition, blockade by the PI3K-specific inhibitor LY294002, Akt inhibitor, the NFκB inhibitor BAY 11-7082, the survivin inhibitor Curcumin, or the survivin inhibitor YM155 reduced the effects of Id1 transfection. These results suggest that the Id1/PI3K/Akt/NFκB/survivin signalling pathway plays a critical role in EPC proliferation. The Id1/PI3K/Akt/NFκB/survivin signalling pathway may represent a novel therapeutic target in the prevention of restenosis after vascular injury.

  14. Antagonism of EGFR and HER3 Enhances the Response to Inhibitors of the PI3K-Akt Pathway in Triple-Negative Breast Cancer

    PubMed Central

    Tao, Jessica J.; Castel, Pau; Radosevic-Robin, Nina; Elkabets, Moshe; Auricchio, Neil; Aceto, Nicola; Weitsman, Gregory; Barber, Paul; Vojnovic, Borivoj; Ellis, Haley; Morse, Natasha; Viola-Villegas, Nerissa Therese; Bosch, Ana; Juric, Dejan; Hazra, Saswati; Singh, Sharat; Kim, Phillip; Bergamaschi, Anna; Maheswaran, Shyamala; Ng, Tony; Penault-Llorca, Frédérique; Lewis, Jason S.; Carey, Lisa A.; Perou, Charles M.; Baselga, José; Scaltriti, Maurizio

    2014-01-01

    Both abundant epidermal growth factor receptor (EGFR or ErbB1) and high activity of the phosphatidyl-inositol 3-kinase (PI3K)–Akt pathway are common and therapeutically targeted in triple-negative breast cancer (TNBC). However, activation of another EGFR family member [human epidermal growth factor receptor 3 (HER3) (or ErbB3)] may limit the antitumor effects of these drugs. We found that TNBC cell lines cultured with the EGFR or HER3 ligand EGF or heregulin, respectively, and treated with either an Akt inhibitor (GDC-0068) or a PI3K inhibitor (GDC-0941) had increased abundance and phosphorylation of HER3. The phosphorylation of HER3 and EGFR in response to these treatments was reduced by the addition of a dual EGFR and HER3 inhibitor (MEHD7945A). MEHD7945A also decreased the phosphorylation (and activation) of EGFR and HER3 and the phosphorylation of downstream targets that occurred in response to the combination of EGFR ligands and PI3K-Akt pathway inhibitors. In culture, inhibition of the PI3K-Akt pathway combined with either MEHD7945A or knockdown of HER3 decreased cell proliferation compared with inhibition of the PI3K-Akt pathway alone. Combining either GDC-0068 or GDC-0941 with MEHD7945A inhibited the growth of xenografts derived from TNBC cell lines or from TNBC patient tumors, and this combination treatment was also more effective than combining either GDC-0068 or GDC-0941 with cetuximab, an EGFR-targeted antibody. After therapy with EGFR-targeted antibodies, some patients had residual tumors with increased HER3 abundance and EGFR/HER3 dimerization (an activating interaction). Thus, we propose that concomitant blockade of EGFR, HER3, and the PI3K-Akt pathway in TNBC should be investigated in the clinical setting. PMID:24667376

  15. Apigenin induces apoptosis through proteasomal degradation of HER2/neu in HER2/neu-overexpressing breast cancer cells via the phosphatidylinositol 3-kinase/Akt-dependent pathway.

    PubMed

    Way, Tzong-Der; Kao, Ming-Ching; Lin, Jen-Kun

    2004-02-06

    Apigenin is a low toxicity and non-mutagenic phytopolyphenol and protein kinase inhibitor. It exhibits anti-proliferating effects on human breast cancer cells. Here we examined several human breast cancer cell lines having different levels of HER2/neu expression and found that apigenin exhibited potent growth-inhibitory activity in HER2/neu-overexpressing breast cancer cells but was much less effective for those cells expressing basal levels of HER2/neu. Induction of apoptosis was also observed in HER2/neu-overexpressing breast cancer cells in a dose- and time-dependent manner. However, the one or more molecular mechanisms of apigenin-induced apoptosis in HER2/neu-overexpressing breast cancer cells remained to be elucidated. A cell survival pathway involving phosphatidylinositol 3-kinase (PI3K), and Akt is known to play an important role in inhibiting apoptosis in response to HER2/neu-overexpressing breast cancer cells, which prompted us to investigate whether this pathway plays a role in apigenin-induced apoptosis in HER2/neu-overexpressing breast cancer cells. Our results showed that apigenin inhibits Akt function in tumor cells in a complex manner. First, apigenin directly inhibited the PI3K activity while indirectly inhibiting the Akt kinase activity. Second, inhibition of HER2/neu autophosphorylation and transphosphorylation resulting from depleting HER2/neu protein in vivo was also observed. In addition, apigenin inhibited Akt kinase activity by preventing the docking of PI3K to HER2/HER3 heterodimers. Therefore, we proposed that apigenin-induced cellular effects result from loss of HER2/neu and HER3 expression with subsequent inactivation of PI3K and AKT in cells that are dependent on this pathway for cell proliferation and inhibition of apoptosis. This implies that the inhibition of the HER2/HER3 heterodimer function provided an especially effective strategy for blocking the HER2/neu-mediated transformation of breast cancer cells. Our results also

  16. Group IB secretory phospholipase A2 promotes matrix metalloproteinase-2-mediated cell migration via the phosphatidylinositol 3-kinase and Akt pathway.

    PubMed

    Choi, Young-Ae; Lim, Hyung-Kyu; Kim, Jae-Ryong; Lee, Chu-Hee; Kim, Young-Jo; Kang, Shin-Sung; Baek, Suk-Hwan

    2004-08-27

    Secretory phospholipase A(2) (sPLA(2)), abundantly expressed in various cells including fibroblasts, is able to promote proliferation and migration. Degradation of collagenous extracellular matrix by matrix metalloproteinase (MMP) plays a role in the pathogenesis of various destructive disorders, such as rheumatoid arthritis, tumor invasion, and metastasis. Here we show that group IB PLA(2) increased pro-MMP-2 activation in NIH3T3 fibroblasts. MMP-2 activity was stimulated by group IB PLA(2) in a dose- and time-dependent manner. Consistent with MMP-2 activation, sPLA(2) decreased expression of type IV collagen. These effects are due to the reduction of tissue inhibitor of metalloproteinase-2 (TIMP-2) and the activation of the membrane type1-MMP (MT1-MMP). The decrease of TIMP-2 levels in conditioned media and the increase of MT1-MMP levels in plasma membrane were observed. In addition, treatment of cells with decanoyl Arg-Val-Lys-Arg-chloromethyl ketone, an inhibitor of pro-MT1-MMP, suppressed sPLA(2)-mediated MMP-2 activation, whereas treatment with bafilomycin A1, an inhibitor of H(+)-ATPase, sustained MMP-2 activation by sPLA(2). The involvement of phosphatidylinositol 3-kinase (PI3K) and Akt in the regulation of MMP-2 activity was further suggested by the findings that PI3K and Akt were phosphorylated by sPLA(2). Expression of p85alpha and Akt mutants, or pretreatment of cells with LY294002, a PI3K inhibitor, attenuated sPLA(2)-induced MMP-2 activation and migration. Taken together, these results suggest that sPLA(2) increases the pro-MMP-2 activation and migration of fibroblasts via the PI3K and Akt-dependent pathway. Because MMP-2 is an important factor directly involved in the control of cell migration and the turnover of extracellular matrix, our study may provide a mechanism for sPLA(2)-promoted fibroblasts migration.

  17. PI3K inhibitors as potential therapeutics for autoimmune disease.

    PubMed

    Ball, Jennifer; Archer, Sophie; Ward, Stephen

    2014-08-01

    Aberrant overactivation of the immune system can give rise to chronic and persistent self-attack, culminating in autoimmune disease. This is currently managed therapeutically using potent immunosuppressive and anti-inflammatory drugs. Class I phosphoinositide 3-kinases (PI3Ks) have been identified as ideal therapeutic targets for autoimmune diseases given their wide-ranging roles in immunological processes. Recent studies into the function of selective PI3K inhibitors in vitro and in vivo have yielded encouraging results, allowing progression into the clinic. Here, we review their recent progress across a range of autoimmune diseases.

  18. INPP4B and PTEN Loss Leads to PI-3,4-P2 Accumulation and Inhibition of PI3K in TNBC.

    PubMed

    Reed, Darien E; Shokat, Kevan M

    2017-02-14

    Triple-negative breast cancer [TNBC, lacks expression of estrogen receptor (ER), progesterone receptor (PR) and amplification of HER2/Neu] remains one of the most aggressive subtypes, affects the youngest patients and still lacks an effective targeted therapy(1,2). Both phosphatidylinositol-3-kinase (PI3K)-α and -β contribute to oncogenesis of solid tumors, including the development of breast cancer(3). Inositol polyphosphate-4-phosphatase type II (INPP4B) catalyzes the removal of the 4'-phosphate of phosphatidylinositol-(3,4-bisphosphate (PI-3,4-P2) creating phosphatidylinositol-3-phosphate(4). There is debate concerning whether PI-3,4-P2 contributes to Akt and downstream effector activation with the known canonical signaling second messenger, phosphatidylinositol-(3,4,5)-trisphosphate (PIP3) (5-7). If PI-3,4-P2 is a positive effector, INPP4B would be a negative regulator of PI3K signaling and there is some evidence to support this(4,8). Utilizing phosphatase and tensin homolog deleted on chromosome ten (PTEN)-null triple-negative breast tumor cell lines, it was unexpectedly found that silencing INPP4B decreased basal phospho-Akt (pAkt) and cellular proliferation, and in most cases sensitized cells to PI3K-α and -β isoform-specific inhibitors. Conversely, overexpression of INPP4B desensitized cells to PI3K inhibitors in a phosphatase activity-dependent manner. In summary, the current investigation of INPP4B in PTEN-null TNBC suggests new mechanistic insight and the potential for targeted therapy for this aggressive subset of breast cancer.

  19. Ursolic Acid Increases Glucose Uptake through the PI3K Signaling Pathway in Adipocytes

    PubMed Central

    He, Yonghan; Li, Wen; Li, Ying; Zhang, Shuocheng; Wang, Yanwen; Sun, Changhao

    2014-01-01

    Background Ursolic acid (UA), a triterpenoid compound, is reported to have a glucose-lowering effect. However, the mechanisms are not fully understood. Adipose tissue is one of peripheral tissues that collectively control the circulating glucose levels. Objective The objective of the present study was to determine the effect and further the mechanism of action of UA in adipocytes. Methods and Results The 3T3-L1 preadipocytes were induced to differentiate and treated with different concentrations of UA. NBD-fluorescent glucose was used as the tracer to measure glucose uptake and Western blotting used to determine the expression and activity of proteins involved in glucose transport. It was found that 2.5, 5 and 10 µM of UA promoted glucose uptake in a dose-dependent manner (17%, 29% and 35%, respectively). 10 µM UA-induced glucose uptake with insulin stimulation was completely blocked by the phosphatidylinositol (PI) 3-kinase (PI3K) inhibitor wortmannin (1 µM), but not by SB203580 (10 µM), the inhibitor of mitogen-activated protein kinase (MAPK), or compound C (2.5 µM), the inhibitor of AMP-activated kinase (AMPK) inhibitor. Furthmore, the downstream protein activities of the PI3K pathway, phosphoinositide-dependent kinase (PDK) and phosphoinositide-dependent serine/threoninekinase (AKT) were increased by 10 µM of UA in the presence of insulin. Interestingly, the activity of AS160 and protein kinase C (PKC) and the expression of glucose transporter 4 (GLUT4) were stimulated by 10 µM of UA under either the basal or insulin-stimulated status. Moreover, the translocation of GLUT4 from cytoplasm to cell membrane was increased by UA but decreased when the PI3K inhibitor was applied. Conclusions Our results suggest that UA stimulates glucose uptake in 3T3-L1 adipocytes through the PI3K pathway, providing important information regarding the mechanism of action of UA for its anti-diabetic effect. PMID:25329874

  20. PREX2 promotes the proliferation, invasion and migration of pancreatic cancer cells by modulating the PI3K signaling pathway

    PubMed Central

    Yang, Jianyi; Gong, Xuejun; Ouyang, Lu; He, Wen; Xiao, Rou; Tan, Li

    2016-01-01

    Phosphatidylinositol-3,4,5-trisphosphate-dependent Rac exchanger factor 2 (PREX2) is a novel regulator of the small guanosine triphosphatase Rac, and has been observed to be implicated in human cancer by inhibiting the activity of phosphatase and tensin homolog (PTEN), thus upregulating the activity of the phosphoinositide 3-kinase (PI3K) signaling pathway. However, the exact role of PREX2 in pancreatic cancer has not been reported to date. In the present study, the expression levels of PREX2 were observed to be frequently increased in pancreatic cancer specimens compared with those in their matched adjacent normal tissues. In addition, PREX2 expression was also frequently upregulated in several pancreatic cancer cell lines, including AsPC-1, BxPC-3, PANC-1 and CFAPC-1, compared with that in the normal pancreatic epithelial cell line HPC-Y5. Overexpression of PREX2 significantly promoted the proliferation, invasion and migration of pancreatic cancer PANC-1 cells, while small interfering RNA-induced knockdown of PREX2 expression significantly inhibited the proliferation, invasion and migration of these cells. Investigation of the molecular mechanism revealed that the overexpression of PREX2 upregulated the phosphorylation levels of PTEN, indicating that the activity of PTEN was reduced, which further increased the phosphorylation levels of AKT, which indicated that the activity of the PI3K signaling pathway was upregulated. By contrast, knockdown of PREX2 upregulated the activity of PTEN and inhibited the activity of the PI3K signaling pathway. In conclusion, the present study demonstrated that PREX2 regulates the proliferation, invasion and migration of pancreatic cancer cells, probably at least via modulation of the activity of PTEN and the PI3K signaling pathway. PMID:27446408

  1. Fruit extracts of Momordica charantia potentiate glucose uptake and up-regulate Glut-4, PPAR gamma and PI3K.

    PubMed

    Kumar, Ramadhar; Balaji, S; Uma, T S; Sehgal, P K

    2009-12-10

    Momordica charantia fruit is a widely used traditional medicinal herb as, anti-diabetic, anti-HIV, anti-ulcer, anti-inflammatory, anti-leukemic, anti-microbial, and anti-tumor. The present study is undertaken to investigate the possible mode of action of fruit extracts derived from Momordica charantia (MC) and study its pharmacological effects for controlling diabetic mellitus. Effects of aqueous and chloroform extracts of Momordica charantia fruit on glucose uptake and up-regulation of glucose transporter (Glut-4), peroxisome proliferator activator receptor gamma (PPAR gamma) and phosphatidylinositol-3 kinase (PI3K), were investigated to show its efficacy as a hypoglycaemic agent. Dose dependent glucose uptake assay was performed on L6 myotubes using 2-deoxy-D-[1-(3)H] glucose. Up-regulatory effects of the extracts on the mRNA expression level of Glut-4, PPAR gamma and PI3K have been studied. The association of Momordica charantia with the aqueous and chloroform extracts of Momordica charantia fruit at 6 microg/ml has shown significant up-regulatory effect, respectively, by 3.6-, 2.8- and 3.8-fold on the battery of targets Glut-4, PPAR gamma and PI3K involved in glucose transport. The up-regulation of glucose uptake was comparable with insulin and rosiglitazone which was approximately 2-fold over the control. Moreover, the inhibitory effect of the cyclohexamide on Momordica charantia fruit extract mediated glucose uptake suggested the requirement of new protein synthesis for the enhanced glucose uptake. This study demonstrated the significance of Glut-4, PPAR gamma and PI3K up-regulation by Momordica charantia in augmenting the glucose uptake and homeostasis.

  2. Insulin/PI3K signaling protects dentate neurons from oxygen-glucose deprivation in organotypic slice cultures.

    PubMed

    Sun, Xiaolu; Yao, Hang; Douglas, Robert M; Gu, Xiang Q; Wang, Juan; Haddad, Gabriel G

    2010-01-01

    It is known that ischemia/reperfusion induces neurodegeneration in the hippocampus in a subregion-dependent manner. This study investigated the mechanism of selective resistance/vulnerability to oxygen-glucose deprivation (OGD) using mouse organotypic hippocampal cultures. Analysis of propidium iodide uptake showed that OGD-induced duration- and subregion-dependent neuronal injury. When compared with the CA1-3 subregions, dentate neuronal survival was more sensitive to inhibition of phosphatidylinositol 3-kinase (PI3K)/Akt signaling under basal conditions. Dentate neuronal sensitivity to PI3K/Akt signaling activation was inversely related to its vulnerability to OGD-induced injury; insulin/insulin-like growth factor 1 pre-treatment conferred neuroprotection to dentate neurons via activation of PI3K/Akt signaling. In contrast, CA1 and CA3 neurons were less sensitive to disruptions of endogenous PI3K/Akt signaling and protective effects of insulin/insulin-like growth factor 1, but more vulnerable to OGD. OGD-induced injury in CA1 was reduced by inhibition of NMDA receptor or mitogen-activated protein kinase signaling, and was prevented by blocking NMDA receptor in the presence of insulin. The CA2 subregion was distinctive in its response to glutamate, OGD, and insulin, compared with other CA subregions. CA2 neurons were sensitive to the protective effects of insulin against OGD-induced injury, but more resistant to glutamate. Distinctive distribution of insulin receptor beta and basal phospho-Akt was detected in our slice cultures. Our results suggest a role for insulin signaling in subregional resistance/vulnerability to cerebral ischemia.

  3. Gefitinib induces lung cancer cell autophagy and apoptosis via blockade of the PI3K/AKT/mTOR pathway

    PubMed Central

    ZHAO, ZHONG-QUAN; YU, ZHONG-YANG; LI, JIE; OUYANG, XUE-NONG

    2016-01-01

    Gefitinib is a selective inhibitor of the tyrosine kinase epidermal growth factor receptor, which inhibits tumor pathogenesis, metastasis and angiogenesis, as well as promoting apoptosis. Therefore, gefitinib presents an effective drug for the targeted therapy of lung cancer. However, the underlying mechanisms by which gefitinib induces lung cancer cell death remain unclear. To investigate the effects of gefitinib on lung cancer cells and the mechanism of such, the present study analyzed the effect of gefitinib on the autophagy, apoptosis and proliferation of the A549 and A549-gefitinib-resistant (GR) cell lines GR. The regulation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/ mammalian target of rapamycin (mTOR) pathway was also investigated. Acridine orange staining revealed that gefitinib induced autophagy of A549 cells but not A549-GR cells. In addition, gefitinib promoted apoptosis and inhibited proliferation of A549 cells but not A549-GR cells. Furthermore, western blot analysis demonstrated that gefitinib treatment led to the downregulation of PI3K, AKT, pAKT, mTOR and phosphorylated-mTOR protein expression in A549 cells but not A549-GR cells. LY294002 blocked the PI3K/AKT/mTOR pathway and induced autophagy and apoptosis of A549 cells, however, no synergistic effect was observed following combined treatment with gefitinib and LY294002. In conclusion, the results of the present study indicate that gefitinib promotes autophagy and apoptosis of lung cancer cells via blockade of the PI3K/AKT/mTOR pathway, which leads to lung cancer cell death. PMID:27347100

  4. Klebsiella pneumoniae Translocates across the Intestinal Epithelium via Rho GTPase- and Phosphatidylinositol 3-Kinase/Akt-Dependent Cell Invasion

    PubMed Central

    Hsu, Chun-Ru; Pan, Yi-Jiun; Liu, Ju-Yun; Chen, Chun-Tang; Lin, Tzu-Lung

    2014-01-01

    Klebsiella pneumoniae is an important pathogen that causes hospital-acquired septicemia and is associated with the recent emergence of community-acquired pyogenic liver abscess (PLA). Clinical typing suggests that K. pneumoniae infections originate from the gastrointestinal reservoir. However, the underlying mechanism remains unknown. Here, we have sought to determine how K. pneumoniae penetrates the intestinal barrier. We identified that bacteremia and PLA clinical isolates adhered to and invaded intestinal epithelial cells. Internalization of K. pneumoniae in three different human colonic cell lines was visualized by confocal microscopy and three-dimensional (3D) imaging. Using a Transwell system, we demonstrated that these K. pneumoniae isolates translocated across a polarized Caco-2 monolayer. No disruptions of transepithelial electrical resistance and altered distribution of tight junction protein ZO-1 or occludin were observed. Therefore, K. pneumoniae appeared to penetrate the intestinal epithelium via a transcellular pathway. Using specific inhibitors, we characterized the host signaling pathways involved. Inhibition by cytochalasin D and nocodazole suggested that actin and microtubule cytoskeleton were both important for K. pneumoniae invasion. A Rho inhibitor, ML141, LY294002, and an Akt1/2 inhibitor diminished K. pneumoniae invasion in a dose-dependent manner, indicating that Rho family GTPases and phosphatidylinositol 3-kinase (PI3K)/Akt signaling were required. By a mouse model of gastrointestinal colonization, in vivo invasion of K. pneumoniae into colonic epithelial cells was demonstrated. Our results present evidence to describe a possible mechanism of gastrointestinal translocation for K. pneumoniae. Cell invasion by manipulating host machinery provides a pathway for gut-colonized K. pneumoniae cells to penetrate the intestinal barrier and access extraintestinal locations to cause disease. PMID:25452552

  5. Klebsiella pneumoniae translocates across the intestinal epithelium via Rho GTPase- and phosphatidylinositol 3-kinase/Akt-dependent cell invasion.

    PubMed

    Hsu, Chun-Ru; Pan, Yi-Jiun; Liu, Ju-Yun; Chen, Chun-Tang; Lin, Tzu-Lung; Wang, Jin-Town

    2015-02-01

    Klebsiella pneumoniae is an important pathogen that causes hospital-acquired septicemia and is associated with the recent emergence of community-acquired pyogenic liver abscess (PLA). Clinical typing suggests that K. pneumoniae infections originate from the gastrointestinal reservoir. However, the underlying mechanism remains unknown. Here, we have sought to determine how K. pneumoniae penetrates the intestinal barrier. We identified that bacteremia and PLA clinical isolates adhered to and invaded intestinal epithelial cells. Internalization of K. pneumoniae in three different human colonic cell lines was visualized by confocal microscopy and three-dimensional (3D) imaging. Using a Transwell system, we demonstrated that these K. pneumoniae isolates translocated across a polarized Caco-2 monolayer. No disruptions of transepithelial electrical resistance and altered distribution of tight junction protein ZO-1 or occludin were observed. Therefore, K. pneumoniae appeared to penetrate the intestinal epithelium via a transcellular pathway. Using specific inhibitors, we characterized the host signaling pathways involved. Inhibition by cytochalasin D and nocodazole suggested that actin and microtubule cytoskeleton were both important for K. pneumoniae invasion. A Rho inhibitor, ML141, LY294002, and an Akt1/2 inhibitor diminished K. pneumoniae invasion in a dose-dependent manner, indicating that Rho family GTPases and phosphatidylinositol 3-kinase (PI3K)/Akt signaling were required. By a mouse model of gastrointestinal colonization, in vivo invasion of K. pneumoniae into colonic epithelial cells was demonstrated. Our results present evidence to describe a possible mechanism of gastrointestinal translocation for K. pneumoniae. Cell invasion by manipulating host machinery provides a pathway for gut-colonized K. pneumoniae cells to penetrate the intestinal barrier and access extraintestinal locations to cause disease. Copyright © 2015, American Society for Microbiology. All

  6. The Prolyl Peptidases PRCP/PREP Regulate IRS-1 Stability Critical for Rapamycin-induced Feedback Activation of PI3K and AKT*

    PubMed Central

    Duan, Lei; Ying, Guoguang; Danzer, Brian; Perez, Ricardo E.; Shariat-Madar, Zia; Levenson, Victor V.; Maki, Carl G.

    2014-01-01

    The phosphatidylinositol 3-kinase (PI3K)/protein kinase B (PKB/AKT)/mammalian target of rapamycin (mTOR) pathway conveys signals from receptor tyrosine kinases (RTKs) to regulate cell metabolism, proliferation, survival, and motility. Previously we found that prolylcarboxypeptidase (PRCP) regulate proliferation and survival in breast cancer cells. In this study, we found that PRCP and the related family member prolylendopeptidase (PREP) are essential for proliferation and survival of pancreatic cancer cells. Depletion/inhibition of PRCP and PREP-induced serine phosphorylation and degradation of IRS-1, leading to inactivation of the cellular PI3K and AKT. Notably, depletion/inhibition of PRCP/PREP destabilized IRS-1 in the cells treated with rapamycin, blocking the feedback activation PI3K/AKT. Consequently, inhibition of PRCP/PREP enhanced rapamycin-induced cytotoxicity. Thus, we have identified PRCP and PREP as a stabilizer of IRS-1 which is critical for PI3K/AKT/mTOR signaling in pancreatic cancer cells. PMID:24936056

  7. RSC96 Schwann Cell Proliferation and Survival Induced by Dilong through PI3K/Akt Signaling Mediated by IGF-I

    PubMed Central

    Chang, Yung-Ming; Kuo, Wu-Hsien; Lai, Tung-Yuan; Shih, Ying-Ting; Tsai, Fuu-Jen; Tsai, Chang-Hai; Shu, Wen-Tong; Chen, Ying-Yu; Chen, Yueh-Sheng; Kuo, Wei-Wen; Huang, Chih-Yang

    2011-01-01

    Schwann cell proliferation is critical for the regeneration of injured nerves. Dilongs are widely used in Chinese herbal medicine to remove stasis and stimulate wound-healing functions. Exactly how this Chinese herbal medicine promotes tissue survival remains unclear. The aim of the present study was to investigate the molecular mechanisms by which Dilong promote neuron regeneration. Our results show that treatment with extract of Dilong induces the phosphorylation of the insulin-like growth factor-I (IGF-I)-mediated phosphatidylinositol 3-kinase/serine-threonine kinase (PI3K/Akt) pathway, and activates protein expression of cell nuclear antigen (PCNA) in a time-dependent manner. Cell cycle analysis showed that G1 transits into the S phase in 12–16 h, and S transits into the G2 phase 20 h after exposure to earthworm extract. Strong expression of cyclin D1, cyclin E and cyclin A occurs in a time-dependent manner. Small interfering RNA (siRNA)-mediated knockdown of PI3K significantly reduced PI3K protein expression levels, resulting in Bcl2 survival factor reduction and a marked blockage of G1 to S transition in proliferating cells. These results demonstrate that Dilong promotes the proliferation and survival of RSC96 cells via IGF-I signaling. The mechanism is mainly dependent on the PI3K protein. PMID:20040524

  8. Dual-Blocking of PI3K and mTOR Improves Chemotherapeutic Effects on SW620 Human Colorectal Cancer Stem Cells by Inducing Differentiation

    PubMed Central

    Kim, Buyun

    2016-01-01

    Cancer stem cells (CSCs) have tumor initiation, self-renewal, metastasis and chemo-resistance properties in various tumors including colorectal cancer. Targeting of CSCs may be essential to prevent relapse of tumors after chemotherapy. Phosphatidylinositol-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) signals are central regulators of cell growth, proliferation, differentiation, and apoptosis. These pathways are related to colorectal tumorigenesis. This study focused on PI3K and mTOR pathways by inhibition which initiate differentiation of SW620 derived CSCs and investigated its effect on tumor progression. By using rapamycin, LY294002, and NVP-BEZ235, respectively, PI3K and mTOR signals were blocked independently or dually in colorectal CSCs. Colorectal CSCs gained their differentiation property and lost their stemness properties most significantly in dual-blocked CSCs. After treated with anti-cancer drug (paclitaxel) on the differentiated CSCs cell viability, self-renewal ability and differentiation status were analyzed. As a result dual-blocking group has most enhanced sensitivity for anti-cancer drug. Xenograft tumorigenesis assay by using immunodeficiency mice also shows that dual-inhibited group more effectively increased drug sensitivity and suppressed tumor growth compared to single-inhibited groups. Therefore it could have potent anti-cancer effects that dual-blocking of PI3K and mTOR induces differentiation and improves chemotherapeutic effects on SW620 human colorectal CSCs. PMID:26955235

  9. Targeting the PI3K/AKT/mTOR and Raf/MEK/ERK pathways in the treatment of breast cancer.

    PubMed

    Saini, Kamal S; Loi, Sherene; de Azambuja, Evandro; Metzger-Filho, Otto; Saini, Monika Lamba; Ignatiadis, Michail; Dancey, Janet E; Piccart-Gebhart, Martine J

    2013-12-01

    Alterations of signal transduction pathways leading to uncontrolled cellular proliferation, survival, invasion, and metastases are hallmarks of the carcinogenic process. The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) and the Raf/mitogen-activated and extracellular signal-regulated kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling pathways are critical for normal human physiology, and also commonly dysregulated in several human cancers, including breast cancer (BC). In vitro and in vivo data suggest that the PI3K/AKT/mTOR and Raf/MEK/ERK cascades are interconnected with multiple points of convergence, cross-talk, and feedback loops. Raf/MEK/ERK and PI3K/AKT/mTOR pathway mutations may co-exist. Inhibition of one pathway can still result in the maintenance of signaling via the other (reciprocal) pathway. The existence of such "escape" mechanisms implies that dual targeting of these pathways may lead to superior efficacy and better clinical outcome in selected patients. Several clinical trials targeting one or both pathways are already underway in BC patients. The toxicity profile of this novel approach of dual pathway inhibition needs to be closely monitored, given the important physiological role of PI3K/AKT/mTOR and Raf/MEK/ERK signaling. In this article, we present a review of the current relevant pre-clinical and clinical data and discuss the rationale for dual inhibition of these pathways in the treatment of BC patients.

  10. [The influence of the aerobic endurance training on the skeletal muscular mitochondria function and PI3K-Akt protein expression].

    PubMed

    Liu, Shao-dong; Zhang, Yan-qiu; Cao, Jiang

    2016-01-01

    To determine the role of phosphatidylinositol 3-kinase--protein kinase B (PI3K-Akt) signaling pathway in the pro- tective effect of aerobic endurance training on the skeletal muscular mitochondria. Thirty-six rats were randomly divided into three groups( n = 12): control group, aerobic endurance training group and one-time exhaustive group. After the intervention, the quadriceps femoris muscle sample was obtained to detect the mitochondrial membrane potential( MMP), the activities of succinate dehydrogenase (SDH) and cy- tochrome coxidase (COX), and the protein levels of p-PI3K and p-Akt. Compared with the control group, the levels of mitochondrial membrane potential, the activities of succinate dehydrogenase and cytochrome coxidase, and the protein levels of p-PI3K and p-Akt were all significantly decreased in the one-time exhaustive group (P < 0.05). However, all the above was partially reversed in the endurance training group (P < 0.05), and there was no obvious difference with the control group (P > 0.05). Aerobic endurance training plays an important role in the protective effect on the skeletal muscular mitochondria, the mechanism may be related to activation PI3K-Akt signaling pathway.

  11. PI3K/Akt Pathway Contributes to Neurovascular Unit Protection of Xiao-Xu-Ming Decoction against Focal Cerebral Ischemia and Reperfusion Injury in Rats

    PubMed Central

    Xiang, Jun; Zhang, Yong; Wang, Guo-Hua; Bao, Jie; Li, Wen-Wei; Zhang, Wen; Xu, Li-Li; Cai, Ding-Fang

    2013-01-01

    In the present study, we used a focal cerebral ischemia and reperfusion rat model to investigate the protective effects of Xiao-Xu-Ming decoction (XXMD) on neurovascular unit and to examine the role of PI3K (phosphatidylinositol 3-kinase)/Akt pathway in this protection. The cerebral ischemia was induced by 90 min of middle cerebral artery occlusion. Cerebral infarct area was measured by tetrazolium staining, and neurological function was observed at 24 h after reperfusion. DNA fragmentation assay, combined with immunofluorescence, was performed to evaluate apoptosis of neuron, astrocyte, and vascular endothelial cell which constitute neurovascular unit. The expression levels of proteins involved in PI3K/Akt pathway were detected by Western blot. The results showed that XXMD improved neurological function, decreased cerebral infarct area and neuronal damage, and attenuated cellular apoptosis in neurovascular unit, while these effects were abolished by inhibition of PI3K/Akt with LY294002. We also found that XXMD upregulated p-PDKl, p-Akt, and p-GSK3β expression levels, which were partly reversed by LY294002. In addition, the increases of p-PTEN and p-c-Raf expression levels on which LY294002 had no effect were also observed in response to XXMD treatment. The data indicated the protective effects of XXMD on neurovascular unit partly through the activation of PI3K/Akt pathway. PMID:23781261

  12. The role of the PI3K/Akt/mTOR signalling pathway in human cancers induced by infection with human papillomaviruses.

    PubMed

    Zhang, Lifang; Wu, Jianhong; Ling, Ming Tat; Zhao, Liang; Zhao, Kong-Nan

    2015-04-17

    Infection with Human papillomaviruses (HPVs) leads to the development of a wide-range of cancers, accounting for 5% of all human cancers. A prominent example is cervical cancer, one of the leading causes of cancer death in women worldwide. It has been well established that tumor development and progression induced by HPV infection is driven by the sustained expression of two oncogenes E6 and E7. The expression of E6 and E7 not only inhibits the tumor suppressors p53 and Rb, but also alters additional signalling pathways that may be equally important for transformation. Among these pathways, the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signalling cascade plays a very important role in HPV-induced carcinogenesis by acting through multiple cellular and molecular events. In this review, we summarize the frequent amplification of PI3K/Akt/mTOR signals in HPV-induced cancers and discuss how HPV oncogenes E6/E7/E5 activate the PI3K/Akt/mTOR signalling pathway to modulate tumor initiation and progression and affect patient outcome. Improvement of our understanding of the mechanism by which the PI3K/Akt/mTOR signalling pathway contributes to the immortalization and carcinogenesis of HPV-transduced cells will assist in devising novel strategies for preventing and treating HPV-induced cancers.

  13. Exposure to bisphenol A at physiological concentrations observed in Chinese children promotes primordial follicle growth through the PI3K/Akt pathway in an ovarian culture system.

    PubMed

    Zhao, Qian; Ma, Yan; Sun, Ning-Xia; Ye, Chen; Zhang, Qing; Sun, Shu-Han; Xu, Chen; Wang, Fang; Li, Wen

    2014-12-01

    The worldwide increase in the use of bisphenol A (BPA) has resulted in increased human exposure, which could affect human reproductive function. Few studies have investigated the effect of BPA exposure on the primordial follicle pool. In this study, we employed a neonatal ovarian culture system comprising organ obtained from female C57BL/6 pups on postnatal day 4 to assess the effect of BPA on the primordial follicle pool. Ovaries were cultured with BPA (0.1 μM, physiological concentration found in children's blood, and 1 μM, 10 μM) or vehicle for 10 days. Our study revealed that the primary follicle number increased during the early time points (⩽5 days), and we observed a reduction in the primordial follicle pool at a later time point (day 10). This reduction at day 10 was due to increased follicle activation and reduced follicle atresia, as determined by immunohistochemistry for Ki-67 and active caspase-3. Then we examined the phosphatidylinositol-3-kinase (PI3K)/Akt pathway, which is known to be important for early follicle growth. BPA exposure induced the upregulation of the PI3K/Akt pathway, which was reversed by concomitant treatment with PI3K inhibitor. Our results reveal a novel mechanism for BPA-induced primordial follicle activation that involves the PI3K/Akt pathway. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Estrogen increases Nrf2 activity through activation of the PI3K pathway in MCF-7 breast cancer cells

    SciTech Connect

    Wu, Juanjuan; Williams, Devin; Walter, Grant A.; Thompson, Winston E.; Sidell, Neil

    2014-11-01

    The actions of the transcription factor Nuclear factor erythroid 2-related factor (Nrf2) in breast cancer have been shown to include both pro-oncogenic and anti-oncogenic activities which is influenced, at least in part, by the hormonal environment. However, direct regulation of Nrf2 by steroid hormones (estrogen and progesterone) has received only scant attention. Nrf2 is known to be regulated by its cytosolic binding protein, Kelch-like ECH-associated protein 1 (Keap1), and by a Keap1-independent mechanism involving a series of phosphorylation steps mediated by phosphatidylinositol 3-kinase (PI3K) and glycogen synthase kinase 3 beta (GSK3β). Here, we report that estrogen (E2) increases Nrf2 activity in MCF7 breast cancer cells through activation of the PI3K/GSK3β pathway. Utilizing antioxidant response element (ARE)-containing luciferase reporter constructs as read-outs for Nrf2 activity, our data indicated that E2 increased ARE activity >14-fold and enhanced the action of the Nrf2 activators, tertiary butylhydroquinone (tBHQ) and sulforaphane (Sul) 4 to 9 fold compared with cells treated with tBHQ or Sul as single agents. This activity was shown to be an estrogen receptor-mediated phenomenon and was antagonized by progesterone. In addition to its action on the reporter constructs, mRNA and protein levels of heme oxygenase 1, an endogenous target gene of Nrf2, was markedly upregulated by E2 both alone and in combination with tBHQ. Importantly, E2-induced Nrf2 activation was completely suppressed by the PI3K inhibitors LY294002 and Wortmannin while the GSK3β inhibitor CT99021 upregulated Nrf2 activity. Confirmation that E2 was, at least partly, acting through the PI3K/GSK3β pathway was indicated by our finding that E2 increased the phosphorylation status of both GSK3β and Akt, a well-characterized downstream target of PI3K. Together, these results demonstrate a novel mechanism by which E2 can regulate Nrf2 activity in estrogen receptor-positive breast cancer

  15. Beclin 1, an Essential Component and Master Regulator of PI3K-III in Health and Disease

    PubMed Central

    McKnight, Nicole C.; Zhenyu, Yue

    2014-01-01

    Autophagy is a cell ‘self-digestion’ pathway involving the synthesis, trafficking and delivery of autophagosomes to lysosomes for degradation. Beclin 1 is a core component of the class III phosphatidylinositol 3-kinase (PI3K-III) complex, which plays an important role in membrane trafficking and restructuring involved in autophagy, endocytosis, cytokinesis and phagocytosis. To date Beclin 1 has largely been characterized in the context of autophagy; it modulates the lipid kinase activity of PI3K-III catalytic unit VPS34, which generates phosphatidylinositol 3-phosphate (PI(3)P), enabling the recruitment of a number of autophagy proteins involved in the nucleation of the autophagosome. Beclin 1 seems to function as an adaptor for recruiting multiple proteins that modulate VPS34. The recent identification of Beclin 1 protein modifications has shed light on its regulation in autophagy, and the discovery of non-autophagy functions of Beclin 1 has expanded our view of Beclin 1's involvement in tissue homeostasis and human diseases. PMID:24729948

  16. Beclin 1, an Essential Component and Master Regulator of PI3K-III in Health and Disease.

    PubMed

    McKnight, Nicole C; Zhenyu, Yue

    2013-12-01

    Autophagy is a cell 'self-digestion' pathway involving the synthesis, trafficking and delivery of autophagosomes to lysosomes for degradation. Beclin 1 is a core component of the class III phosphatidylinositol 3-kinase (PI3K-III) complex, which plays an important role in membrane trafficking and restructuring involved in autophagy, endocytosis, cytokinesis and phagocytosis. To date Beclin 1 has largely been characterized in the context of autophagy; it modulates the lipid kinase activity of PI3K-III catalytic unit VPS34, which generates phosphatidylinositol 3-phosphate (PI(3)P), enabling the recruitment of a number of autophagy proteins involved in the nucleation of the autophagosome. Beclin 1 seems to function as an adaptor for recruiting multiple proteins that modulate VPS34. The recent identification of Beclin 1 protein modifications has shed light on its regulation in autophagy, and the discovery of non-autophagy functions of Beclin 1 has expanded our view of Beclin 1's involvement in tissue homeostasis and human diseases.

  17. Anaplasma phagocytophilum infection modulates expression of megakaryocyte cell cycle genes through phosphatidylinositol-3-kinase signaling.

    PubMed

    Khanal, Supreet; Sultana, Hameeda; Catravas, John D; Carlyon, Jason A; Neelakanta, Girish

    2017-01-01

    Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis infects neutrophils and other cells from hematopoietic origin. Using human megakaryocytic cell line, MEG-01, we show that expression of cell cycle genes in these cells are altered upon A. phagocytophilum infection. Expression of several cell cycle genes in MEG-01 cells was significantly up regulated at early and then down regulated at later stages of A. phagocytophilum infection. Lactate dehydrogenase (LDH) assays revealed reduced cellular cytotoxicity in MEG-01 cells upon A. phagocytophilum infection. The levels of both PI3KCA (p110 alpha, catalytic subunit) and PI3KR1 (p85, regulatory subunit) of Class I PI3 kinases and phosphorylated protein kinase B (Akt/PKB) and inhibitory kappa B (IκB) were elevated at both early and late stages of A. phagocytophilum infection. Inhibition of PI3 kinases with LY294002 treatment resulted in significant reduction in the expression of tested cell cycle genes, A. phagocytophilum burden and phosphorylated Akt levels in these MEG-01 cells. Collectively, these results suggest a role for PI3K-Akt-NF-κB signaling pathway in the modulation of megakaryocyte cell cycle genes upon A. phagocytophilum infection.

  18. Signals controlling un-differentiated states in embryonic stem and cancer cells: role of the phosphatidylinositol 3' kinase pathway.

    PubMed

    Voskas, Daniel; Ling, Ling Sunny; Woodgett, James Robert

    2014-10-01

    The capacity of embryonic stem (ES) cells to differentiate into cell lineages comprising the three germ layers makes them powerful tools for studying mammalian early embryonic development in vitro. The human body consists of approximately 210 different somatic cell types, the majority of which have limited proliferative capacity. However, both stem cells and cancer cells bypass this replicative barrier and undergo symmetric division indefinitely when cultured under defined conditions. Several signal transduction pathways play important roles in regulating stem cell development, and aberrant expression of components of these pathways is linked to cancer. Among signaling systems, the critical role of leukemia inhibitory factor (LIF) coupled to the Jak/STAT3 (signal transduction and activation of transcription-3) pathway in maintaining stem cell self-renewal has been extensively reviewed. This pathway additionally plays multiple roles in tumorigenesis. Likewise, the phosphatidylinositide 3-kinase (PI3K)/protein kinase B (PKB/Akt) pathway has been determined to play an important role in both stem cell maintenance and tumor development. This pathway is often induced in cancer with frequent mutational activation of the catalytic subunit of PI3K or loss of a primary PI3K antagonist, phosphatase and tensin homolog deleted on chromosome ten (PTEN). This review focusses on roles of the PI3K signal transduction pathway components, with emphasis on functions in stem cell maintenance and cancer. Since the PI3K pathway impinges on and collaborates with other signaling pathways in regulating stem cell development and/or cancer, aspects of the canonical Wnt, Ras/mitogen-activated protein kinase (MAPK), and TGF-β signaling pathways are also discussed.

  19. The therapeutic potential of targeting the PI3K pathway in pediatric brain tumors.

    PubMed

    Rogers, Hazel A; Estranero, Jasper; Gudka, Keshni; Grundy, Richard G

    2017-01-10

    Central nervous system tumors are the most common cancer type in children and the leading cause of cancer related deaths. There is therefore a need to develop novel treatments. Large scale profiling studies have begun to identify alterations that could be targeted therapeutically, including the phosphoinositide 3-kinase (PI3K) signaling pathway, which is one of the most commonly activated pathways in cancer with many inhibitors under clinical development. PI3K signaling has been shown to be aberrantly activated in many pediatric CNS neoplasms. Pre-clinical analysis supports a role for PI3K signaling in the control of tumor growth, survival and migration as well as enhancing the cytotoxic effects of current treatments. Based on this evidence agents targeting PI3K signaling have begun to be tested in clinical trials of pediatric cancer patients. Overall, targeting the PI3K pathway presents as a promising strategy for the treatment of pediatric CNS tumors. In this review we examine the genetic alterations found in the PI3K pathway in pediatric CNS tumors and the pathological role it plays, as well as summarizing the current pre-clinical and clinical data supporting the use of PI3K pathway inhibitors for the treatment of these tumors.

  20. Potent efficacy of combined PI3K/mTOR and JAK or ABL inhibition in murine xenograft models of Ph-like acute lymphoblastic leukemia.

    PubMed

    Tasian, Sarah K; Teachey, David T; Li, Yong; Shen, Feng; Harvey, Richard C; Chen, I-Ming; Ryan, Theresa; Vincent, Tiffaney L; Willman, Cheryl L; Perl, Alexander E; Hunger, Stephen P; Loh, Mignon L; Carroll, Martin; Grupp, Stephan A

    2017-01-12

    Philadelphia chromosome (Ph)-like B-cell acute lymphoblastic leukemia (Ph-like ALL) is associated with activated JAK/STAT, Abelson kinase (ABL), and/or phosphatidylinositol 3-kinase (PI3K) signaling and poor clinical outcomes. PI3K pathway signaling inhibitors have been minimally investigated in Ph-like ALL. We hypothesized that targeted inhibition of PI3Kα, PI3Kδ, PI3K/mTOR, or target of rapamycin complex 1/2 (TORC1/TORC2) would decrease leukemia proliferation and abrogate aberrant kinase signaling and that combined PI3K pathway and JAK inhibition or PI3K pathway and SRC/ABL inhibition would have superior efficacy compared to inhibitor monotherapy. We treated 10 childhood ALL patient-derived xenograft models harboring various Ph-like genomic alterations with 4 discrete PI3K pathway protein inhibitors and observed marked leukemia reduction and in vivo signaling inhibition in all models. Treatment with dual PI3K/mTOR inhibitor gedatolisib resulted in near eradication of ALL in cytokine receptor-like factor 2 (CRLF2)/JAK-mutant models with mean 92.2% (range, 86.0%-99.4%) reduction vs vehicle controls (P < .0001) and in prolonged animal survival. Gedatolisib also inhibited ALL proliferation in ABL/platelet-derived growth factor receptor (PDGFR)-mutant models with mean 66.9% (range, 42.0%-87.6%) reduction vs vehicle (P < .0001). Combined gedatolisib and ruxolitinib treatment of CRLF2/JAK-mutant models more effectively inhibited ALL proliferation than either inhibitor alone (P < .001) and further enhanced survival. Similarly, superior efficacy of combined gedatolisib and dasatinib was observed in ABL/PDGFR-mutant models (P < .001). Overall, PI3K/mTOR inhibition potently decreased ALL burden in vivo; antileukemia activity was further enhanced with combination inhibitor therapy. Clinical trials testing combinations of kinase inhibitors in Ph-like ALL patients are indicated. © 2017 by The American Society of Hematology.

  1. c-Met Overexpression Contributes to the Acquired Apoptotic Resistance of Nonadherent Ovarian Cancer Cells through a Cross Talk Mediated by Phosphatidylinositol 3-Kinase and Extracellular Signal-Regulated Kinase 1/212

    PubMed Central

    Tang, Maggie K S; Zhou, Hong Y; Yam, Judy W P; Wong, Alice S T

    2010-01-01

    Ovarian cancer is the most lethal gynecologic cancer mainly because of widespread peritoneal dissemination and malignant ascites. Key to this is the capacity of tumor cells to escape suspension-induced apoptosis (anoikis), which also underlies their resistance to chemotherapy. Here, we used a nonadherent cell culture model to investigate the molecular mechanisms of apoptotic resistance of ovarian cancer cells that may mimic the chemoresistance found in solid tumors. We found that ovarian cancer cells acquired a remarkable resistance to anoikis and apoptosis induced by exposure to clinically relevant doses of two front-line chemotherapeutic drugs cisplatin and paclitaxel when grown in three-dimensional than monolayer cultures. Inhibition of the hepatocyte growth factor (HGF) receptor c-Met, which is frequently overexpressed in ovarian cancer, by a specific inhibitor or small interfering RNA blocked the acquired anoikis resistance and restored chemosensitivity in three-dimensional not in two-dimensional cultures. These effects were found to be dependent on both phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK) 1/2 signaling pathways. Inhibitors of PI3K/Akt abrogated ERK1/2 activation and its associated anoikis resistance in response to HGF, suggesting a signaling relay between these two pathways. Furthermore, we identified a central role of Ras as a mechanism of this cross talk. Interestingly, Ras did not lie upstream of PI3K/Akt, whereas PI3K/Akt signaling to ERK1/2 involved Ras. These findings shed new light on the apoptotic resistance mechanism of nonadherent ovarian cancer ascites cells and may have important clinical implications. PMID:20126471

  2. Daily Exposure to Di(2-ethylhexyl) Phthalate Alters Estrous Cyclicity and Accelerates Primordial Follicle Recruitment Potentially Via Dysregulation of the Phosphatidylinositol 3-Kinase Signaling Pathway in Adult Mice1

    PubMed Central

    Hannon, Patrick R.; Peretz, Jackye; Flaws, Jodi A.

    2014-01-01

    ABSTRACT Humans are exposed daily to di(2-ethylhexyl) phthalate (DEHP), a plasticizer found in many consumer, medical, and building products containing polyvinyl chloride. Large doses of DEHP disrupt normal ovarian function; however, the effects of DEHP at environmentally relevant levels, the effects of DEHP on folliculogenesis, and the mechanisms by which DEHP disrupts ovarian function are unclear. The present study tested the hypothesis that relatively low levels of DEHP disrupt estrous cyclicity as well as accelerate primordial follicle recruitment by dysregulating phosphatidylinositol 3-kinase (PI3K) signaling. Adult CD-1 mice were orally dosed with DEHP (20 μg/kg/day–750 mg/kg/day) daily for 10 and 30 days. Following dosing, the effects on estrous cyclicity were examined, and follicle numbers were histologically quantified. Further, the ovarian mRNA and protein levels of PI3K signaling factors that are associated with early folliculogenesis were quantified. The data indicate that 10- and 30-day exposure to DEHP prolonged the duration of estrus and accelerated primordial follicle recruitment. Specifically, DEHP exposure decreased the percentage of primordial follicles and increased the percentage of primary follicles counted following 10-day exposure and increased the percentage of primary follicles counted following 30-day exposure. DEHP exposure, at doses that accelerate folliculogenesis, increased the levels of 3-phosphoinositide-dependent protein kinase-1, mammalian target of rapamycin complex 1, and protein kinase B and decreased the levels of phosphatase and tensin homolog, potentially driving PI3K signaling. Collectively, relatively low levels of DEHP disrupt estrous cyclicity and accelerate primordial follicle recruitment potentially via a mechanism involving dysregulation of PI3K signaling. PMID:24804967

  3. Effects of rehabilitation training on apoptosis of nerve cells and the recovery of neural and motor functions in rats with ischemic stroke through the PI3K/Akt and Nrf2/ARE signaling pathways.

    PubMed

    Jin, Xiao-Fei; Wang, Shan; Shen, Min; Wen, Xin; Han, Xin-Rui; Wu, Jun-Chang; Tang, Gao-Zhuo; Wu, Dong-Mei; Lu, Jun; Zheng, Yuan-Lin

    2017-09-01

    This study was designed in order to investigate the effects between rehabilitation training on the apoptosis of nerve cells and the recovery of neural and motor functions of rats with ischemic stroke by way of the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) and nuclear factor E2-related factor 2/antioxidant responsive element (Nrf2/ARE) signaling pathways. In total, 110 healthy adult male Sprague-Dawley (SD) rats were selected in order to take part in this study. Ninety SD rats were used in order to establish the middle cerebral artery occlusion (MCAO), among which 80 rats were randomly assigned as part of the natural recovery, natural recovery+Rp-PI3K (the rats injected with PI3K/Akt inhibitor LY294002), rehabilitation training, and rehabilitation training+Rp-PI3K groups. Meanwhile, 20 rats were selected as part of the sham operation group. The neural and motor functions of these rats were evaluated using a balance beam test and the Bederson score. The mRNA expressions of PI3K, Akt, Nrf2 and HO-1 were measured using an RT-qPCR. The protein expressions of PI3K, p-PI3K, Akt, p-Akt, Nrf2 and HO-1 were also detected by using western blotting and the immunohistochemistry process. The cell cycle and cell apoptosis were detected by using a flow cytometry and TUNEL assay. The sham operation group exhibited lower neural and motor function scores than other groups. At the 7, 14, and 21 d marks of this study, the neural and motor function scores were increased in the natural recovery, natural recovery+Rp-PI3K, and rehabilitation training+Rp-PI3K groups in comparison with the rehabilitation training group but found to be decreased in the natural recovery group in comparison with the natural recovery+Rp-PI3K group. In comparison with the sham operation group, expressions of PI3K, Nrf2 and HO-1, and proportions of p-PI3K/PI3K and p-Akt/Akt were all higher in the natural recovery, rehabilitation training, and rehabilitation training+Rp-PI3K groups. Same trends were

  4. Pik3ip1 Modulates Cardiac Hypertrophy by Inhibiting PI3K Pathway

    PubMed Central

    Song, Hong Ki; Kim, Jiyeon; Lee, Jong Sub; Nho, Kyoung Jin; Jeong, Hae Chang; Kim, Jihwa; Ahn, Youngkeun; Park, Woo Jin; Kim, Do Han

    2015-01-01

    Cardiac hypertrophy is an adaptive response to various physiological and pathological stimuli. Phosphoinositide-3 kinase (PI3K) is a highly conserved lipid kinase involved in physiological cardiac hypertrophy (PHH). PI3K interacting protein1 (Pik3ip1) shares homology with the p85 regulatory subunit of PI3K and is known to interact with the p110 catalytic subunit of PI3K, leading to attenuation of PI3K activity in liver and immune cells. However, the role of Pik3ip1 in the heart remains unknown. In the present study, the effects of Pik3ip1 on cardiac hypertrophy were examined. We found that the expression level of Pik3ip1 was markedly higher in cardiomyocytes than in fibroblasts. The interaction of Pik3ip1 with the p110a subunit of PI3K in the heart was identified by immunoprecipitation using neonatal rat cardiomyocytes (NRCM). Approximately 35% knockdown of Pik3ip1 was sufficient to induce myocardial hypertrophy. Pik3ip1 deficiency was shown to lead to activation of PI3K/protein kinase B (AKT)/ mammalian target of rapamycin (mTOR) signaling pathway, increasing protein synthesis and cell size. However, adenovirus-mediated overexpression of Pik3ip1 attenuated PI3K-mediated cardiac hypertrophy. Pik3ip1 was upregulated by PHH due to swimming training, but not by pathological cardiac hypertrophy (PAH) due to pressure-overload, suggesting that Pik3ip1 plays a compensatory negative role for PHH. Collectively, our results elucidate the mechanisms for the roles of Pik3ip1 in PI3K/AKT signaling pathway. PMID:25826393

  5. Blueberry Phytochemicals Inhibit Growth and Metastatic Potential of MDA-MB-231 Breast Cancer Cells Through Modulation of the Phosphatidylinositol 3-Kinase Pathway

    PubMed Central

    Adams, Lynn S.; Phung, Sheryl; Yee, Natalie; Seeram, Navindra P.; Li, Liya; Chen, Shiuan

    2010-01-01

    Dietary phytochemicals are known to exhibit a variety of anti-carcinogenic properties. This study investigated the chemopreventive activity of blueberry extract in triple negative breast cancer cell lines in vitro and in vivo. Blueberry decreased cell proliferation in HCC38, HCC1937 and MDA-MB-231 cells with no effect on the non-tumorigenic MCF-10A cell line. Decreased metastatic potential of MDA-MB-231 cells by blueberry was shown through inhibition of cell motility using wound healing assays and migration through a PET membrane. Blueberry treatment decreased the activity of matrix metalloproteinase 9 and the secretion of urokinase-type plasminogen activator while increasing tissue inhibitor of metalloproteinase-1 and plasminogen activator inhibitor-1 secretion in MDA-MB-231 conditioned medium as shown by western blotting. Cell signaling pathways that control the expression/activation of these processes were investigated via western blotting and reporter gene assay. Treatment with blueberry decreased phosphatidylinositol 3-kinase (PI3K)/AKT and nuclear factor kappa-B (NFκB) activation in MDA-MB-231 cells where protein kinase C (PKC) and extracellular regulated kinase (ERK) were not affected. In vivo, the efficacy of blueberry to inhibit triple negative breast tumor growth was evaluated using the MDA-MB-231 xenograft model. Tumor weight and proliferation (Ki-67 expression) were decreased in blueberry treated mice, where apoptosis (caspase-3 expression) was increased compared to controls. Immunohistochemical analysis of tumors from blueberry-fed mice showed decreased activation of AKT and p65 NFκB signaling proteins with no effect on the phosphorylation of ERK. These data illustrate the inhibitory effect of blueberry phytochemicals on the growth and metastatic potential of MDA-MB-231 cells through modulation of the PI3K/AKT/NFκB pathway. PMID:20388778

  6. Protective effects of statins on L-DOPA neurotoxicity due to the activation of phosphatidylinositol 3-kinase and free radical scavenging in PC12 cell culture.

    PubMed

    Koh, Seong-Ho; Park, Hyun-Hee; Choi, Na-Young; Lee, Kyu-Yong; Kim, Sangjae; Lee, Young Joo; Kim, Hee-Tae

    2011-01-25

    Neurotoxic effects have been suggested for l-3,4-dihydroxyphenylalanine (L-DOPA), while neuroprotective effects have been proposed for statins. We investigated whether certain statins (simvastatin or pitavastatin) could inhibit L-DOPA neurotoxicity. Neuronally-differentiated PC12 (nPC12) cells were treated with L-DOPA and/or statins for 24h, and their viabilities were measured using a cell counting kit, trypan blue staining, and 4',6-diamidino-2-phenylindole (DAPI) staining. Free radical and specific intracellular signaling protein levels were measured with 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) and Western blotting, respectively. High concentrations of l-DOPA reduced nPC12 cell viability, but combined treatment with statins restored viability. Treatment with 200 μM L-DOPA increased free radical and hydroxyl radical levels, but combined treatment with 5 μM statins decreased these levels. Survival-related signaling proteins were decreased in nPC12 cells treated with 200 μM L-DOPA, but combined treatment with 5μM statins significantly increased the levels of these proteins. Treatment with 200 μM L-DOPA significantly increased death-related signaling proteins, while combined treatment with 5 μM statins reduced the levels of these proteins. Pretreatment with LY294002, a phosphatidylinositol 3 kinase (PI3K) inhibitor, before combined treatment with statins and L-DOPA almost completely blocked the protective effects of statins. These results indicate that statins reduce L-DOPA neurotoxicity by lowering oxidative stress and by enhancing survival signals and inhibiting death signals via activation of the PI3K pathway.

  7. The PI3K/Akt/mTOR signaling pathway mediates insulin-like growth factor 1-induced E-cadherin down-regulation and cell proliferation in ovarian cancer cells.

    PubMed

    Lau, Man-Tat; Leung, Peter C K

    2012-12-30

    Insulin-like growth factor 1 (IGF1) is produced by ovarian cancer cells and it has been suggested that it plays an important role in tumor progression. In this study, we report that IGF1 treatment down-regulated E-cadherin by up-regulating E-cadherin transcriptional repressors, Snail and Slug, in human ovarian cancer cells. The pharmacological inhibition of phosphatidylinositol-3-kinase (PI3K) and mammalian target of rapamycin (mTOR) suggests that PI3K/Akt/mTOR signaling is required for IGF1-induced E-cadherin down-regulation. Moreover, IGF1 up-regulated Snail and Slug expression via the PI3K/Akt/mTOR signaling pathway. Finally, IGF1-induced cell proliferation was abolished by inhibiting PI3K/Akt/mTOR signaling. This study demonstrates a novel mechanism in which IGF1 down-regulates E-cadherin expression through the activation of PI3K/Akt/mTOR signaling and the up-regulation of Snail and Slug in human ovarian cancer cells.

  8. PI3K/SHIP2/PTEN pathway in cell polarity and hepatitis C virus pathogenesis

    PubMed Central

    Awad, Aline; Gassama-Diagne, Ama

    2017-01-01

    Hepatitis C virus (HCV) infects hepatocytes, polarized cells in the liver. Chronic HCV infection often leads to steatosis, fibrosis, cirrhosis and hepatocellular carcinoma, and it has been identified as the leading cause of liver transplantation worldwide. The HCV replication cycle is dependent on lipid metabolism and particularly an accumulation of lipid droplets in host cells. Phosphoinositides (PIs) are minor phospholipids enriched in different membranes and their levels are tightly regulated by specific PI kinases and phosphatases. PIs are implicated in a vast array of cellular responses that are central to morphogenesis, such as cytoskeletal changes, cytokinesis and the recruitment of downstream effectors to govern mechanisms involved in polarization and lumen formation. Important reviews of the literature identified phosphatidylinositol (PtdIns) 4-kinases, and their lipid products PtdIns(4)P, as critical regulators of the HCV life cycle. SH2-containing inositol polyphosphate 5-phosphatase (SHIP2), phosphoinositide 3-kinase (PI3K) and their lipid products PtdIns(3,4)P2 and PtdIns(3,4,5)P3, respectively, play an important role in the cell membrane and are key to the establishment of apicobasal polarity and lumen formation. In this review, we will focus on these new functions of PI3K and SHIP2, and their deregulation by HCV, causing a disruption of apicobasal polarity, actin organization and extracellular matrix assembly. Finally we will highlight the involvement of this pathway in the event of insulin resistance and nonalcoholic fatty liver disease related to HCV infection. PMID:28105255

  9. Inhibition of PI3K Prevents the Proliferation and Differentiation of Human Lung Fibroblasts into Myofibroblasts: The Role of Class I P110 Isoforms

    PubMed Central

    Conte, Enrico; Fruciano, Mary; Fagone, Evelina; Gili, Elisa; Caraci, Filippo; Iemmolo, Maria; Crimi, Nunzio; Vancheri, Carlo

    2011-01-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive fibroproliferative disease characterized by an accumulation of fibroblasts and myofibroblasts in the alveolar wall. Even though the pathogenesis of this fatal disorder remains unclear, transforming growth factor-β (TGF-β)-induced differentiation and proliferation of myofibroblasts is recognized as a primary event. The molecular pathways involved in TGF-β signalling are generally Smad-dependent yet Smad-independent pathways, including phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt), have been recently proposed. In this research we established ex-vivo cultures of human lung fibroblasts and we investigated the role of the PI3K/Akt pathway in two critical stages of the fibrotic process induced by TGF-β: fibroblast proliferation and differentiation into myofibroblasts. Here we show that the pan-inhibitor of PI3Ks LY294002 is able to abrogate the TGF-β-induced increase in cell proliferation, in α- smooth muscle actin expression and in collagen production besides inhibiting Akt phosphorylation, thus demonstrating the centrality of the PI3K/Akt pathway in lung fibroblast proliferation and differentiation. Moreover, for the first time we show that PI3K p110δ and p110γ are functionally expressed in human lung fibroblasts, in addition to the ubiquitously expressed p110α and β. Finally, results obtained with both selective inhibitors and gene knocking-down experiments demonstrate a major role of p110γ and p110α in both TGF-β-induced fibroblast proliferation and differentiation. This finding suggests that specific PI3K isoforms can be pharmacological targets in IPF. PMID:21984893

  10. PTEN regulates apoptotic cell death through PI3-K/Akt/GSK3β signaling pathway in DMH induced early colon carcinogenesis in rat.

    PubMed

    Saini, Manpreet Kaur; Sanyal, Sankar Nath

    2012-08-01

    Phosphatidylinositol 3-kinase (PI3-K) and Akt (protein kinase B), are both essential signaling molecules that are up-regulated in various cancers. Here, we examined the molecular mechanisms by which PI3-K and Akt expression are regulated by glycogen synthase kinase-3β (GSK-3β) and the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in the early stages of experimental colon carcinogenesis. 1,2-dimethylhydrazine (DMH) was utilized for the induction of colon cancer while piroxicam, a traditional non-steroidal anti-inflammatory drug and c-phycocyanin, a biliprotein from Spirulina platensis (cyanobacterium) as the chemopreventive agents. Western blotting and immunofluorescence results indicated that the expression of PI3-K and Akt was promoted in the DMH group while least apoptosis was detected in this group as analyzed by Hoechst 33342-propidium iodide co-staining. DMH group further detected lower GSK-3β and PTEN expression as compared to other groups. Piroxicam and c-phycocyanin treatment resulted significant apoptotic cell death while showing low PI3-K and Akt expressions. Mitochondrial membrane potential (ΔΨ(M)) alterations (examined by JC-1 and rhodamine 123 labeling of colonocytes) and fluorescence intensity measurement of ROS level, were also analyzed showing the raised ΔΨ(M) while reduced ROS levels in DMH group, however piroxicam and c-phycocyanin treatment resulted in falling of ΔΨ(M) although both stimulated the ROS production as analyzed by flow cytometry. The present study thus identified that piroxicam, a traditional NSAID and c-phycocyanin, a newly discovered COX-2 selective inhibitor, constitute remarkable chemopreventive targets in mediating apoptosis in the DMH induced early rat colon carcinogenesis via regulating PI3-K/Akt/GSK-3β/PTEN signaling pathways. Further, a combination of the two drugs provides a better therapeutic option, than the monotherapy regimen.

  11. Psoralidin inhibits proliferation and enhances apoptosis of human esophageal carcinoma cells via NF-κB and PI3K/Akt signaling pathways

    PubMed Central

    Jin, Zhiliang; Yan, Wei; Jin, Hui; Ge, Changzheng; Xu, Yanhua

    2016-01-01

    Esophageal cancer is the most common gastrointestinal cancer. Psoralidin exhibits antioxidant, anti-apoptotic, anti-inflammatory and antitumor effects, which result in the inhibition of cancer formation. The present study aimed to investigate the effect of psoralidin on esophageal carcinoma proliferation and growth, and to elucidate its underlying mechanism of action. The effect of psoralidin on cell proliferation was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Using an annexin V-fluorescein isothiocyanate/propidium iodide apoptosis detection kit and 4′,6-diamidino-2-phenylindole staining assay, the present study demonstrated that psoralidin significantly enhanced apoptosis of human esophageal carcinoma Eca9706 cells. In addition, caspase-3 activity was analyzed with a caspase-3 colorimetric assay kit, while nuclear factor (NF)-κB activity and protein phosphatidylinositol 3-kinase (PI3K)/Akt expression were measured with an NF-κB enzyme-linked immunosorbent assay kit and western blot analysis, respectively. Eca9706 cells were treated with a PI3K agonist in order to investigate the mechanism of action of psoralidin. It was observed that psoralidin was able to decrease the proliferation and promote the cellular apoptosis of Eca9706 cells in a dose-dependent manner. Furthermore, psoralidin was also able to inhibit the caspase-3 activity of Eca9706 cells in a dose-dependent manner. In addition, psoralidin inhibited NF-κB activity and reduced PI3K and Akt protein expression in Eca9706 cells. Notably, the PI3K agonist was able to reverse the effect of psoralidin on Eca9706 cells. The results of the present study demonstrated that psoralidin was able to inhibit proliferation and enhance apoptosis of human esophageal carcinoma cells via the NF-κB and PI3K/Akt signaling pathways. PMID:27446379

  12. Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways.

    PubMed

    Yu, Jian-Bo; Shi, Jia; Zhang, Yuan; Gong, Li-Rong; Dong, Shu-An; Cao, Xin-Shun; Wu, Li-Li; Wu, Li-Na

    2015-01-01

    Electroacupuncture at select acupoints have been verified to protect against organ dysfunctions during endotoxic shock. And, heme oxygenase (HO)-1 as a phase II enzyme and antioxidant contributed to the protection of kidney in septic shock rats. The phosphatidylinositol 3-kinase (PI3K)-Akt pathway mediated the activation of NF-E2 related factor-2 (Nrf2), which was involved in HO-1 induction. To understand the efficacy of electroacupuncture stimulation in ameliorating acute kidney injury (AKI) through the PI3K/Akt/Nrf2 pathway and subsequent HO-1 upregulation, a dose of LPS 5mg/kg was administered intravenously to replicate the rabbit model of AKI induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Neiguan acupoints for five consecutive days while sham electroacupuncture at non-acupoints as control. Results displayed that electroacupuncture stimulation significantly alleviated the morphologic renal damage, attenuated renal tubular apoptosis, suppressed the elevated biochemical indicators of AKI caused by LPS, enhanced the expressions of phospho-Akt, HO-1protein, Nrf2 total and nucleoprotein, and highlighted the proportions of Nrf2 nucleoprotein as a parallel. Furthermore, partial protective effects of elecroacupuncture were counteracted by preconditioning with wortmannin (the selective PI3K inhibitor), indicating a direct involvement of PI3K/Akt pathway. Inconsistently, wortmannin pretreatment made little difference to the expressions of HO-1, Nrf2 nucleoprotein and total protein, which indicated that PI3K/Akt may be not the only pathway responsible for electroacupuncture-afforded protection against LPS-induced AKI. These findings provide new insights into the potential future clinical applications of electroacupuncture for AKI induced by endotoxic shock instead of traditional remedies.

  13. Upregulated WDR26 serves as a scaffold to coordinate PI3K/ AKT pathway-driven breast cancer cell growth, migration, and invasion.

    PubMed

    Ye, Yuanchao; Tang, Xiaoyun; Sun, Zhizeng; Chen, Songhai

    2016-04-05

    The phosphatidylinositol 3-kinase (PI3K)/AKT pathway transmits signals downstream of receptor tyrosine kinases and G protein-coupled receptors (GPCRs), and is one of the most dysregulated pathways in breast cancer. PI3Ks and AKTs consist of multiple isoforms that play distinct and even opposite roles in breast cancer cell growth and metastasis. However, it remains unknown how the activities of various PI3K and AKT isoforms are coordinated during breast cancer progression. Previously, we showed WDR26 is a novel WD40 protein that binds Gβγ and promotes Gβγ signaling. Here, we demonstrate that WDR26 is overexpressed in highly malignant breast tumor cell lines and human breast cancer samples, and that WDR26 overexpression correlates with shortened survival of breast cancer patients. In highly malignant cell lines (MDA-MB231, DU4475 and BT549), downregulation of WDR26 expression selectively alleviated GPCR- but not EGF receptor-stimulated PI3K/AKT signaling and tumor cell growth, migration and invasion. In contrast, in a less malignant cell line (MCF7), WDR26 overexpression had the opposite effect. Additional studies indicate that downstream of GPCR stimulation, WDR26 serves as a scaffold that fosters assembly of a specific signaling complex consisting of Gβγ, PI3Kβ and AKT2. In an orthotopic xenograft mouse model of breast cancer, disrupting formation of this complex, by overexpressing WDR26 mutants in MDA-MB231 cells, abrogated PI3K/AKT activation and tumor cell growth and metastasis. Together, our results identify a novel mechanism regulating GPCR-dependent activation of the PI3K/AKT signaling axis in breast tumor cells, and pinpoint WDR26 as a potential therapeutic target for breast cancer.

  14. Hydrostatic Compress Force Enhances the Viability and Decreases the Apoptosis of Condylar Chondrocytes through Integrin-FAK-ERK/PI3K Pathway

    PubMed Central

    Ma, Dandan; Kou, Xiaoxing; Jin, Jing; Xu, Taotao; Wu, Mengjie; Deng, Liquan; Fu, Lusi; Liu, Yi; Wu, Gang; Lu, Haiping

    2016-01-01

    Reduced mechanical stimuli in many pathological cases, such as hemimastication and limited masticatory movements, can significantly affect the metabolic activity of mandibular condylar chondrocytes and the growth of mandibles. However, the molecular mechanisms for these phenomena remain unclear. In this study, we hypothesized that integrin-focal adhesion kinase (FAK)-ERK (extracellular signal–regulated kinase)/PI3K (phosphatidylinositol-3-kinase) signaling pathway mediated the cellular response of condylar chondrocytes to mechanical loading. Primary condylar chondrocytes were exposed to hydrostatic compressive forces (HCFs) of different magnitudes (0, 50, 100, 150, 200, and 250 kPa) for 2 h. We measured the viability, morphology, and apoptosis of the chondrocytes with different treatments as well as the gene, protein expression, and phosphorylation of mechanosensitivity-related molecules, such as integrin α2, integrin α5, integrin β1, FAK, ERK, and PI3K. HCFs could significantly increase the viability and surface area of condylar chondrocytes and decrease their apoptosis in a dose-dependent manner. HCF of 250 kPa resulted in a 1.51 ± 0.02-fold increase of cell viability and reduced the ratio of apoptotic cells from 18.10% ± 0.56% to 7.30% ± 1.43%. HCFs could significantly enhance the mRNA and protein expression of integrin α2, integrin α5, and integrin β1 in a dose-dependent manner, but not ERK1, ERK2, or PI3K. Instead, HCF could significantly increase phosphorylation levels of FAK, ERK1/2, and PI3K in a dose-dependent manner. Cilengitide, the potent integrin inhibitor, could dose-dependently block such effects of HCFs. HCFs enhances the viability and decreases the apoptosis of condylar chondrocytes through the integrin-FAK-ERK/PI3K pathway. PMID:27827993

  15. Id-1 promotes tumorigenicity and metastasis of human esophageal cancer cells through activation of PI3K/AKT signaling pathway.

    PubMed

    Li, Bin; Tsao, Sai Wah; Li, Yuk Yin; Wang, Xianghong; Ling, Ming Tat; Wong, Yong Chuan; He, Qing Yu; Cheung, Annie L M

    2009-12-01

    Id-1 (inhibitor of differentiation or DNA binding) is a helix-loop-helix protein that is overexpressed in many types of cancer including esophageal squamous cell carcinoma (ESCC). We previously reported that ectopic Id-1 expression activates the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) signaling pathway in human esophageal cancer cells. In this study, we confirmed a positive correlation between Id-1 and phospho-AKT (Ser473) expressions in ESCC cell lines, as well as in ESCC on a tissue microarray. To investigate the significance of Id-1 in esophageal cancer progression, ESCC cells with stable ectopic Id-1 expression were inoculated subcutaneously into the flank of nude mice and were found to form larger tumors that showed elevated Ki-67 proliferation index and increased angiogenesis, as well as reduced apoptosis, compared with control cells expressing the empty vector.The Id-1-overexpressing cells also exhibited enhanced metastatic potential in the experimental metastasis assay. Treatment with the PI3K inhibitor LY294002 attenuated the tumor promotion effects of Id-1, indicating that the effects were mediated by the PI3K/AKT signaling pathway. In addition, our in vitro experiments showed that ectopic Id-1 expression altered the expression levels of markers associated with epithelial-mesenchymal transition and enhanced the migration ability of esophageal cancer cells. The Id-1-overexpressing ESCC cells also exhibited increased invasive potential, which was in part due to PI3K/AKT-dependent modulation of matrix metalloproteinase-9 expression. In conclusion, our results provide the first evidence that Id-1 promotes tumorigenicity and metastasis of human esophageal cancer in vivo and that the PI3K inhibitor LY294002 can attenuate these effects.

  16. PI3K/Akt is involved in brown adipogenesis mediated by growth differentiation factor-5 in association with activation of the Smad pathway

    SciTech Connect

    Hinoi, Eiichi; Iezaki, Takashi; Fujita, Hiroyuki; Watanabe, Takumi; Odaka, Yoshiaki; Ozaki, Kakeru; Yoneda, Yukio

    2014-07-18

    Highlights: • Akt is preferentially phosphorylated in BAT and sWAT of aP2-GDF5 mice. • PI3K/Akt signaling is involved in GDF5-induced brown adipogenesis. • PI3K/Akt signaling regulates GDF5-induced Smad5 phosphorylation. - Abstract: We have previously demonstrated promotion by growth differentiation factor-5 (GDF5) of brown adipogenesis for systemic energy expenditure through a mechanism relevant to activating the bone morphological protein (BMP) receptor/mothers against decapentaplegic homolog (Smad)/peroxisome proliferator-activated receptor gamma co-activator 1α (PGC-1α) pathway. Here, we show the involvement of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in brown adipogenesis mediated by GDF5. Overexpression of GDF5 in cells expressing adipocyte protein-2 markedly accelerated the phosphorylation of Smad1/5/8 and Akt in white and brown adipose tissues. In brown adipose tissue from heterozygous GDF5{sup Rgsc451} mutant mice expressing a dominant-negative (DN) GDF5 under obesogenic conditions, the basal phosphorylation of Smad1/5/8 and Akt was significantly attenuated. Exposure to GDF5 not only promoted the phosphorylation of both Smad1/5/8 and Akt in cultured brown pre-adipocytes, but also up-regulated Pgc1a and uncoupling protein-1 expression in a manner sensitive to the PI3K/Akt inhibitor Ly294002 as well as retroviral infection with DN-Akt. GDF5 drastically promoted BMP-responsive luciferase reporter activity in a Ly294002-sensitive fashion. Both Ly294002 and DN-Akt markedly inhibited phosphorylation of Smad5 in the nuclei of brown pre-adipocytes. These results suggest that PI3K/Akt signals play a role in the GDF5-mediated brown adipogenesis through a mechanism related to activation of the Smad pathway.

  17. Inhibition of infarction-induced sympathetic innervation with endothelin receptor antagonism via a PI3K/GSK-3β-dependent pathway.

    PubMed

    Lee, T-M; Chang, Nen-Chung; Lin, Shinn-Zong

    2017-03-01

    Although endothelin (ET)-1 has been shown to upregulate nerve growth factor (NGF) expression, the molecular mechanisms are largely unknown. Phosphatidylinositol 3-kinase (PI3K)/Akt/glycogen synthase kinase (GSK)-3β signal has been implicated in the regulation of NGF. We investigated whether selective ET receptor blockers attenuated cardiac sympathetic reinnervation through restoring PI3K/Akt/GSK-3β activity. After ligation of the left anterior descending artery, male Wistar rats were randomized to either vehicle, atrasentan (an ETA receptor antagonist) or A-192621 (an ETB receptor antagonist) for 4 weeks. Sympathetic hyperinnervation after infarction was confirmed by myocardial norepinephrine measurement and immunofluorescent analysis. Post infarction was associated with increased reactive oxygen species (ROS), as measured by myocardial superoxide levels and dihydroethidine fluorescence staining. This was paralleled by a significant upregulation of NGF expression on mRNA and protein levels in the vehicle-treated rats, which reduced after administering atrasentan, not A-192621. Arrhythmic scores in the vehicle-treated rats were significantly higher than those treated with atrasentan. In an in vivo study atrasentan-induced decreased NGF was associated with activation of PI3K/Akt signaling pathway, which was further confirmed by the ex vivo study showing the restoration of NGF levels after coadministration of PI3K inhibitors (wortmannin and LY294002). Lithium chloride, an inhibitor of GSK-3β, did not provide additional attenuated NGF levels compared with atrasentan alone. Finally, atrasentan-attenuated NGF levels were reversed in the presence of peroxynitrite generator. ETA receptor antagonism is a mediator to attenuate sympathetic hyperinnervation probably through restoration of PI3K/Akt/GSK-3β/ROS signaling pathway, a potential pharmacological target for arrhythmias after infarction.

  18. Electroacupuncture Ameliorates Acute Renal Injury in Lipopolysaccharide-Stimulated Rabbits via Induction of HO-1 through the PI3K/Akt/Nrf2 Pathways

    PubMed Central

    Gong, Li-rong; Dong, Shu-an; Cao, Xin-shun; Wu, Li-li; Wu, Li-na

    2015-01-01

    Electroacupuncture at select acupoints have been verified to protect against organ dysfunctions during endotoxic shock. And, heme oxygenase (HO)-1 as a phase II enzyme and antioxidant contributed to the protection of kidney in septic shock rats. The phosphatidylinositol 3-kinase (PI3K)-Akt pathway mediated the activation of NF-E2 related factor-2 (Nrf2), which was involved in HO-1 induction. To understand the efficacy of electroacupuncture stimulation in ameliorating acute kidney injury (AKI) through the PI3K/Akt/Nrf2 pathway and subsequent HO-1 upregulation, a dose of LPS 5mg/kg was administered intravenously to replicate the rabbit model of AKI induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Neiguan acupoints for five consecutive days while sham electroacupuncture at non-acupoints as control. Results displayed that electroacupuncture stimulation significantly alleviated the morphologic renal damage, attenuated renal tubular apoptosis, suppressed the elevated biochemical indicators of AKI caused by LPS, enhanced the expressions of phospho-Akt, HO-1protein, Nrf2 total and nucleoprotein, and highlighted the proportions of Nrf2 nucleoprotein as a parallel. Furthermore, partial protective effects of elecroacupuncture were counteracted by preconditioning with wortmannin (the selective PI3K inhibitor), indicating a direct involvement of PI3K/Akt pathway. Inconsistently, wortmannin pretreatment made little difference to the expressions of HO-1, Nrf2 nucleoprotein and total protein, which indicated that PI3K/Akt may be not the only pathway responsible for electroacupuncture-afforded protection against LPS-induced AKI. These findings provide new insights into the potential future clinical applications of electroacupuncture for AKI induced by endotoxic shock instead of traditional remedies. PMID:26524181

  19. The Chinese herbal medicine FTZ attenuates insulin resistance via IRS1 and PI3K in vitro and in rats with metabolic syndrome.

    PubMed

    Hu, Xuguang; Wang, Man; Bei, Weijian; Han, Zongyu; Guo, Jiao

    2014-02-20

    Insulin resistance plays an important role in the development of metabolic syndrome (MS). Fu Fang Zhen Zhu Tiao Zhi formula (FTZ), a Chinese medicinal decoction, has been used to relieve hyperlipidemia, atherosclerosis and other symptoms associated with metabolic disorders in the clinic. To evaluate the effect of FTZ on insulin resistance, HepG2 cells were induced with high insulin as a model of insulin resistance and treated with FTZ at one of three dosages. Next, the levels of glucose content, insulin receptor substrate1 (IRS1) protein expression and phosphatidylinositol 3-kinase (PI3K) subunit p85 mRNA expression were measured. Alternatively, MS was induced in rats via gavage feeding of a high-fat diet for four consecutive weeks followed by administration of FTZ for eight consecutive weeks. Body weight and the plasma levels of lipids, insulin and glucose were evaluated. Finally, the expression of PI3K p85 mRNA in adipose tissue of rats was measured. Our results revealed that FTZ attenuated glucose content and up-regulated the expression of PI3K p85 mRNA and IRS1 protein in insulin-resistant HepG2 cells in vitro. Moreover, FTZ reduced body weight and the plasma concentrations of triacylglycerol, cholesterol, fasting glucose and insulin in insulin resistant MS rats. FTZ also elevated the expression of PI3K p85 mRNA in the adipose tissues of MS rats. FTZ attenuated MS symptoms by decreasing the plasma levels of glucose and lipids. The underlying mechanism was attenuation of the reduced expression of PI3K p85 mRNA and IRS1 protein in both insulin-resistant HepG2 cells and MS rats.

  20. The Chinese herbal medicine FTZ attenuates insulin resistance via IRS1 and PI3K in vitro and in rats with metabolic syndrome

    PubMed Central

    2014-01-01

    Background Insulin resistance plays an important role in the development of metabolic syndrome (MS). Fu Fang Zhen Zhu Tiao Zhi formula (FTZ), a Chinese medicinal decoction, has been used to relieve hyperlipidemia, atherosclerosis and other symptoms associated with metabolic disorders in the clinic. Methods To evaluate the effect of FTZ on insulin resistance, HepG2 cells were induced with high insulin as a model of insulin resistance and treated with FTZ at one of three dosages. Next, the levels of glucose content, insulin receptor substrate1 (IRS1) protein expression and phosphatidylinositol 3-kinase (PI3K) subunit p85 mRNA expression were measured. Alternatively, MS was induced in rats via gavage feeding of a high-fat diet for four consecutive weeks followed by administration of FTZ for eight consecutive weeks. Body weight and the plasma levels of lipids, insulin and glucose were evaluated. Finally, the expression of PI3K p85 mRNA in adipose tissue of rats was measured. Results Our results revealed that FTZ attenuated glucose content and up-regulated the expression of PI3K p85 mRNA and IRS1 protein in insulin-resistant HepG2 cells in vitro. Moreover, FTZ reduced body weight and the plasma concentrations of triacylglycerol, cholesterol, fasting glucose and insulin in insulin resistant MS rats. FTZ also elevated the expression of PI3K p85 mRNA in the adipose tissues of MS rats. Conclusion FTZ attenuated MS symptoms by decreasing the plasma levels of glucose and lipids. The underlying mechanism was attenuation of the reduced expression of PI3K p85 mRNA and IRS1 protein in both insulin-resistant HepG2 cells and MS rats. PMID:24555840

  1. Combretastatin A4 Regulates Proliferation, Migration, Invasion, and Apoptosis of Thyroid Cancer Cells via PI3K/Akt Signaling Pathway

    PubMed Central

    Liang, Weixin; Lai, Yongqiang; Zhu, Mingzhang; Huang, Shangshu; Feng, Weizhao; Gu, Xiaoyu

    2016-01-01

    Background Combretastatin A4 (CA4) is a potential therapeutic candidate for a variety of human cancer treatments. However, the inhibitive effects of CA4 on thyroid cancer cells are still not well-clarified. This study aimed to investigate the potential effect of CA4 on thyroid cancer cells, as well as underlying mechanism. Material/Methods Human thyroid papillary carcinoma cell line TPC1 was pre-treated with 5 concentrations of CA4 (0, 1, 2, 5, or 10 μM) for 2 h. Cell proliferation was determined by 3-(4, 5-dimethyl-2- thiazolyl)-2, 5-diphenyl -2-H-tetrazolium bromide (MTT) assay. Cell migration and invasion were detected by a modified Boyden chamber assay. Moreover, cell apoptosis was detected by terminal deoxynucleotidyl (TUNEL) staining assay and flow cytometry method. Western blot analysis was performed to determine the expression changes of epithelial-mesenchymal transition (EMT)-related proteins and phosphatidylinositol-3-kinase/serine/threonine kinase (PI3K/Akt) signaling pathway proteins. Results CA4 significantly inhibited the cell proliferation, migration, and invasion, and significantly promoted cell apoptosis in a dose-dependent manner compared with the control group. The EMT-related protein levels of N-Cadherin, Vimentin, Snail1, Slug, Twist1, and ZEB1 were significantly decreased by CA4, while E-cadherin had no significant difference compared with the control group. Moreover, PI3K/Akt signaling pathway protein levels of p-PI3K and p-Akt were significantly decreased, whereas PI3K and Akt had no significant differences compared with the control group. Conclusions CA4 can inhibit proliferation, migration, and invasion and promote apoptosis of TPC1 cells. These effects might be through the PI3K/Akt signaling pathway. CA4 may be a potential therapeutic target for the treatment of thyroid cancer. PMID:27966519

  2. Computational studies of Ras and PI3K

    NASA Technical Reports Server (NTRS)

    Ren, Lei; Cucinotta, Francis A.

    2004-01-01

    Until recently, experimental techniques in molecular cell biology have been the primary means to investigate biological risk upon space radiation. However, computational modeling provides an alternative theoretical approach, which utilizes various computational tools to simulate proteins, nucleotides, and their interactions. In this study, we are focused on using molecular mechanics (MM) and molecular dynamics (MD) to study the mechanism of protein-protein binding and to estimate the binding free energy between proteins. Ras is a key element in a variety of cell processes, and its activation of phosphoinositide 3-kinase (PI3K) is important for survival of transformed cells. Different computational approaches for this particular study are presented to calculate the solvation energies and binding free energies of H-Ras and PI3K. The goal of this study is to establish computational methods to investigate the roles of different proteins played in the cellular responses to space radiation, including modification of protein function through gene mutation, and to support the studies in molecular cell biology and theoretical kinetics models for our risk assessment project.

  3. Physical association of PDK1 with AKT1 is sufficient for pathway activation independent of membrane localization and phosphatidylinositol 3 kinase.

    PubMed

    Ding, Zhiyong; Liang, Jiyong; Li, Jin; Lu, Yiling; Ariyaratna, Vathsala; Lu, Zhimin; Davies, Michael A; Westwick, John K; Mills, Gordon B

    2010-03-26

    Frequent activation of the AKT serine-threonine kinase in cancer confers resistance to therapy. AKT is activated by a multi-step process involving phosphatidylinositide (PtdIns) phosphate-mediated recruitment of AKT and its upstream kinases, including 3-Phosphoinositide-dependent kinase 1 (PDK1), to the inner surface of the cell membrane. PDK1 in the appropriate context phosphorylates AKT at threonine 308 (T308) to activate AKT. Whether PtdIns(3,4,5)Ps (PtdInsP3) binding and AKT membrane translocation mediate functions other than formation of a functional PDK1::AKT complex have not been fully elucidated. We fused complementary fragments of intensely fluorescent protein (IFP) to AKT1 and PDK1 to induce a stable complex to study the prerequisites of AKT1 phosphorylation and function. In the stabilized PDK1-IFPC::IFPN-AKT1 complex, AKT1 T308 phosphorylation was independent of PtdIns, as demonstrated by treatment with Phosphatidylinositol 3 Kinase (PI3K) inhibitors. Further when interaction with PtdIns and the cell membrane was prevented by creating PH-domain mutants of AKT1 (R25A) and PDK1 (R474A), AKT1 phosphorylation on T308 was maintained in the PDK1-IFPC::IFPN-AKT1 complex. The PDK1-IFPC::IFPN-AKT1 complex was sufficient for phosphorylation of known AKT substrates, and conferred resistance to inhibitors of PI3K (LY294002, PI103, GDC0941 and TGX286) but not inhibitors of the downstream TORC1 complex (rapamycin). Thus the locus of action of targeted therapeutics can be elucidated by the constitutively active AKT1 complex. Our data indicate that PtdIns and membrane localization are not required for AKT phosphorylation and activation, but rather serve to induce a functional physical interaction between PDK1 and AKT. The PDK1-IFPC::IFPN-AKT1 complex provides a cell-based platform to examine specificity of drugs targeting PI3K pathway components.

  4. Expression quantitative trait loci for PI3K/AKT pathway

    PubMed Central

    Ryu, Dongchan; Lee, Chaeyoung

    2017-01-01

    Abstract A genome-wide association study (GWAS) was conducted to identify expression quantitative trait loci (eQTLs) for the genes involved in phosphatidylinositol-3-kinase/v-akt murine thymoma viral oncogene homolog (PI3K/AKT) pathway. Data on mRNA expression of 341 genes in lymphoblastoid cell lines of 373 Europeans recruited by the 1000 Genomes Project using Illumina HiSeq2000 were utilized. We used their genotypes at 5,941,815 nucleotide variants obtained by Genome Analyzer II and SOLiD. The association analysis revealed 4166 nucleotide variants associated with expression of 85 genes (P < 5 × 10−8). A total of 73 eQTLs were identified as association signals for the expression of multiple genes. They included 9 eQTLs for both of the genes encoding collagen type I alpha 1 (COL1A1) and integrin alpha 11 (ITGA11), which synthesize a major complex of plasma membrane. They also included eQTLs for type IV collagen molecules; 13 eQTLs for both collagen type IV alpha 1 (COL4A1) and collagen type IV alpha 2 (COL4A2) and 18 eQTLs for both collagen type IV alpha 5 (COL4A5) and collagen type IV alpha 6 (COL4A6). Some genes expressed by the eQTLs might induce expression of the genes encoding type IV collagen. One eQTL (rs16871986) was located in the promoter of palladin (PALLD) gene which might synthesize collagen by activating fibroblasts through the PI3K/AKT pathway. Another eQTL (rs34845474) was located in an enhancer of cadherin related family member 3 (CDHR3) gene which can mediate cell adhesion. This study showed a profile of eQTLs for the genes involved in the PI3K/AKT pathway using a healthy population, revealing 73 eQTLs associated with expression of multiple genes. They might be candidates of common variants in predicting genetic susceptibility to cancer and in targeting cancer therapy. Further studies are required to examine their underlying mechanisms for regulating expression of the genes. PMID:28072738

  5. Skeletal muscle PI3K p110β regulates expression of AMP-activated protein kinase.

    PubMed

    Matheny, Ronald W; Abdalla, Mary N; Geddis, Alyssa V; Leandry, Luis A; Lynch, Christine M

    2017-01-22

    Skeletal muscle metabolic homeostasis is maintained through numerous biochemical and physiological processes. Two principal molecular regulators of skeletal muscle metabolism include AMP-activated protein kinase (AMPK) and phosphatidylinositol 3-kinase (PI3K); however, PI3K exists as multiple isoforms, and specific metabolic actions of each isoform have not yet been fully elucidated in skeletal muscle. Given this lack of knowledge, we performed a series of experiments to define the extent to which PI3K p110β mediated expression and (or) activation of AMPK in skeletal muscle. To determine the effect of p110β inhibition on AMPK expression and phosphorylation in cultured cells, C2C12 myoblasts were treated with a pharmacological inhibitor of p110β (TGX-221), siRNA against p110β, or overexpression of kinase-dead p110β. Expression and phosphorylation of AMPK were unaffected in myoblasts treated with TGX-221 or expressing kinase-dead p110β. However, expressions of total and phosphorylated AMPK at T172 were reduced in myoblasts treated with p110β siRNA. When normalized to expression of total AMPK, phosphorylation of AMPK S485/491 was elevated in p110β-deficient myoblasts. Similar results were observed in tibialis anterior muscle from mice with conditional deletion of p110β (p110β-mKO mice). Analysis of AMPK transcript expression revealed decreased expression of Prkaa2 in p110β-deficient myoblasts and in p110β-mKO muscle. Loss of p110β had no effect on oligomycin-stimulated phosphorylation of AMPK or phosphorylated Acetyl-CoA carboxylase (ACC), although oligomycin-induced AMPK and ACC phosphorylation were increased in p110β-deficient myoblasts compared to oligomycin-stimulated control myoblasts when normalized to levels of total AMPK or ACC. Overall, these results suggest that p110β positively regulates expression of AMPK in cultured myoblasts and in skeletal muscle in vivo; moreover, despite the reduced abundance of AMPK in p110β-deficient myoblasts, loss

  6. Astaxanthin reduces isoflurane-induced neuroapoptosis via the PI3K/Akt pathway.

    PubMed

    Wang, Chun-Mei; Cai, Xiao-Lan; Wen, Qing-Ping

    2016-05-01

    Astaxanthin is an oxygen-containing derivative of carotenoids that effectively suppresses reactive oxygen and has nutritional and medicinal value. The mechanisms underlying the effects of astaxanthin on isoflurane‑induced neuroapoptosis remain to be fully understood. The present study was conducted to evaluate the protective effect of astaxanthin to reduce isoflurane‑induced neuroapoptosis and to investigate the underlying mechanisms. The results demonstrated that isoflurane induced brain damage, increased caspase‑3 activity and suppressed the phosphatidylinositol 3‑kinase (PI3K)/protein kinase B (Akt) signaling pathway in an in vivo model. However, treatment with astaxanthin significantly inhibited brain damage, suppressed caspase‑3 activity and upregulated the PI3K/Akt pathway in the isoflurane‑induced rats. Furthermore, isoflurane suppressed cell growth, induced cell apoptosis, enhanced caspase‑3 activity and downregulated the PI3K/Akt pathway in organotypic hippocampal slice culture. Administration of astaxanthin significantly promoted cell growth, reduced cell apoptosis and caspase‑3 activity, and upregulated the PI3K/Akt pathway and isoflurane‑induced neuroapoptosis. The present study demonstrated that downregulation of the PI3K/Akt pathway reduced the effect of astaxanthin to protect against isoflurane‑induced neuroapoptosis in the in vitro model. The results of the current study suggested that the protective effect of astaxanthin reduces the isoflurane-induced neuroapoptosis via activation of the PI3K/Akt signaling pathway.

  7. Mice Expressing Activated PI3K Rapidly Develop Advanced Colon Cancer

    PubMed Central

    Leystra, Alyssa A.; Deming, Dustin A.; Zahm, Christopher D.; Farhoud, Mohammed; Paul Olson, Terrah J.; Hadac, Jamie N.; Nettekoven, Laura A.; Albrecht, Dawn M.; Clipson, Linda; Sullivan, Ruth; Washington, Mary Kay; Torrealba, Jose R.; Weichert, Jamey P.; Halberg, Richard B.

    2012-01-01

    Aberrations in the phosphatidylinositide-3-kinase (PI3K) signaling pathway play a key role in the pathogenesis of numerous cancers by altering cellular growth, metabolism, proliferation, and apoptosis (1). Mutations in the catalytic domain of PI3K that generate a dominantly active kinase are commonly found in human colorectal cancers and have been thought to drive tumor progression, but not initiation (2). However, the effects of constitutively activated PI3K upon the intestinal mucosa have not been previously studied in animal models. Here, we demonstrate that the expression of a dominantly active form of the PI3K protein in the mouse intestine results in hyperplasia and advanced neoplasia. Mice expressing constitutively active PI3K in the epithelial cells of the distal small bowel and colon rapidly developed invasive adenocarcinomas in the colon that spread into the mesentery and adjacent organs. The histological characteristics of these tumors were strikingly similar to invasive mucinous colon cancers in humans. Interestingly, these tumors formed without a benign polypoid intermediary, consistent with the lack of aberrant WNT signaling observed. Together, our findings indicate a non-canonical mechanism of colon tumor initiation that is mediated through activation of PI3K. This unique model has the potential to further our understanding of human disease and facilitate the development of therapeutics through pharmacologic screening and biomarker identification. PMID:22525701

  8. Nerve Growth Factor Protects the Ischemic Heart via Attenuation of the Endoplasmic Reticulum Stress Induced Apoptosis by Activation of Phosphatidylinositol 3-Kinase

    PubMed Central

    Wei, Ke; Liu, Li; Xie, Fei; Hao, Xuechao; Luo, Jie; Min, Su

    2015-01-01

    Background: Increased expression of nerve growth factor (NGF) has been found in the myocardium suffered from ischemia and reperfusion (I/R). The pro-survival activity of NGF on ischemic heart has been supposed to be mediated by phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway. Endoplasmic reticulum (ER) stress, which is activated initially as a defensive response to eliminate the accumulated unfolded proteins, has shown a critical involvement in the ischemia induced myocardial apoptosis. This study was aimed to investigate whether NGF induced heart protection against I/R injury includes a mechanism of attenuation of ER stress-induced myocardial apoptosis by activation of PI3K/Akt pathway. Methods: Isolated adult rat hearts were perfused with a Langendörff perfusion system. Hearts in the Sham group were subjected to 225 min of continuous Krebs-Henseleit buffer (KHB) perfusion without ischemia. Hearts in I/R group were perfused with KHB for a 75-min of equilibration period followed by 30 min of global ischemia and 120 min of KHB reperfusion. Hearts in the NGF group accepted 45 min of euilibration perfusion and 30 min of NGF pretreatment (with a final concentration of 100 ng/ml in the KHB) before 30 min of global ischemia and 120 min of reperfusion. Hearts in K252a and LY294002 groups were pretreated with either a TrkA inhibitor, K252a or a phosphatidyl inositol 3-kinase inhibitor, LY294002 for 30 min before NGF (100 ng/ml) administration. Cardiac hemodynamics were measured from the beginning of the perfusion. Cardiac enzymes and cardiac troponin I (cTnI) were assayed before ischemia and at the end of reperfusion. Myocardial apoptosis rate was measured by TUNEL staining, and expression of glucose-related protein 78 (GRP78), CCAAT/enhancer-binding protein homologous protein (CHOP), caspase-12, total- and phospho-(Ser473)-Akt were assessed by Western blot analyses. Results: NGF pretreatment significantly improved the recovery of post

  9. Nerve growth factor protects the ischemic heart via attenuation of the endoplasmic reticulum stress induced apoptosis by activation of phosphatidylinositol 3-kinase.

    PubMed

    Wei, Ke; Liu, Li; Xie, Fei; Hao, Xuechao; Luo, Jie; Min, Su

    2015-01-01

    Increased expression of nerve growth factor (NGF) has been found in the myocardium suffered from ischemia and reperfusion (I/R). The pro-survival activity of NGF on ischemic heart has been supposed to be mediated by phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway. Endoplasmic reticulum (ER) stress, which is activated initially as a defensive response to eliminate the accumulated unfolded proteins, has shown a critical involvement in the ischemia induced myocardial apoptosis. This study was aimed to investigate whether NGF induced heart protection against I/R injury includes a mechanism of attenuation of ER stress-induced myocardial apoptosis by activation of PI3K/Akt pathway. Isolated adult rat hearts were perfused with a Langendörff perfusion system. Hearts in the Sham group were subjected to 225 min of continuous Krebs-Henseleit buffer (KHB) perfusion without ischemia. Hearts in I/R group were perfused with KHB for a 75-min of equilibration period followed by 30 min of global ischemia and 120 min of KHB reperfusion. Hearts in the NGF group accepted 45 min of euilibration perfusion and 30 min of NGF pretreatment (with a final concentration of 100 ng/ml in the KHB) before 30 min of global ischemia and 120 min of reperfusion. Hearts in K252a and LY294002 groups were pretreated with either a TrkA inhibitor, K252a or a phosphatidyl inositol 3-kinase inhibitor, LY294002 for 30 min before NGF (100 ng/ml) administration. Cardiac hemodynamics were measured from the beginning of the perfusion. Cardiac enzymes and cardiac troponin I (cTnI) were assayed before ischemia and at the end of reperfusion. Myocardial apoptosis rate was measured by TUNEL staining, and expression of glucose-related protein 78 (GRP78), CCAAT/enhancer-binding protein homologous protein (CHOP), caspase-12, total- and phospho-(Ser473)-Akt were assessed by Western blot analyses. NGF pretreatment significantly improved the recovery of post-ischemia cardiac hemodynamics. Reduced

  10. Muc-1 promotes migration and invasion of oral squamous cell carcinoma cells via PI3K-Akt signaling.

    PubMed

    Li, Ping; Xiao, Li Ying; Tan, Hong

    2015-01-01

    Muc-1 is a member of the carbohydrate-binding protein family that contributes to neoplastic transformation, tumor survival, angiogenesis, and metastasis. The aim of this study is to investigate the role of muc-1 in human oral squamous cell carcinoma progression. In this study, we tested our hypothesis that muc-1 regulate oral squamous cell carcinoma cells (SCC-9) malignant biological behaviors, and silencing muc-1 reduced SCC-9 cellular colony forming ability, migration and invasion. Moreover, silenced cells present defects in phosphatidylinositol 3-kinase (PI3K)-serine/threonine kinase (Akt) signaling, and reduced expression/activity of matrix metallopeptidase (MMP)-2/9. Furthermore, in muc-1 siRNA-transfected cells, we detected a decrease in signal transducer and activator of transcription 3 (STAT3) phosphorylation and nuclear translocation. In vivo, muc-1 siRNA cells inoculated subcutaneously in nude mice demonstrated decreased tumor growth and PI3K-Akt signaling inhibition. These results indicate that muc-1 is a key factor in SCC-9 tumor migration, invasion, and suggesting that muc-1 can be a novel therapeutic target in oral squamous cell carcinoma.

  11. Nonsteroidal anti-inflammatory drugs modulate cellular glycosaminoglycan synthesis by affecting EGFR and PI3K signaling pathways

    PubMed Central

    Mozolewski, Paweł; Moskot, Marta; Jakóbkiewicz-Banecka, Joanna; Węgrzyn, Grzegorz; Bocheńska, Katarzyna; Banecki, Bogdan; Gabig-Cimińska, Magdalena

    2017-01-01

    In this report, selected non-steroidal anti-inflammatory drugs (NSAIDs), indomethacin and nimesulide, and analgesics acetaminophen, alone, as well as in combination with isoflavone genistein as potential glycosaminoglycan (GAG) metabolism modulators were considered for the treatment of mucopolysaccharidoses (MPSs) with neurological symptoms due to the effective blood-brain barrier (BBB) penetration properties of these compounds. We found that indomethacin and nimesulide, but not acetaminophen, inhibited GAG synthesis in fibroblasts significantly, while the most pronounced impairment of glycosaminoglycan production was observed after exposure to the mixture of nimesulide and genistein. Phosphorylation of the EGF receptor (EGFR) was inhibited even more effective in the presence of indomethacin and nimesulide than in the presence of genistein. When examined the activity of phosphatidylinositol-3-kinase (PI3K) production, we observed its most significant decrease in the case of fibroblast exposition to nimesulide, and afterwards to indomethacin and genistein mix, rather than indomethacin used alone. Some effects on expression of individual GAG metabolism-related and lysosomal function genes, and significant activity modulation of a number of genes involved in intracellular signal transduction pathways and metabolism of DNA and proteins were detected. This study documents that NSAIDs, and their mixtures with genistein modulate cellular glycosaminoglycan synthesis by affecting EGFR and PI3K signaling pathways. PMID:28240227

  12. Involvement of PI3K and MMP1 in PDGF-induced Migration of Human Adipose-derived Stem Cells.

    PubMed

    Lim, Yoonhwa; Lee, Minji; Jeong, Hyeju; Kim, Haekwon

    2017-06-01

    Human adult stem cells have widely been examined for their clinical application including their wound healing effect in vivo. To function as therapeutic cells, however, cells must represent the ability of directed migration in response to signals. This study aimed to investigate the mechanism of platelet-derived growth factor (PDGF)-induced migration of the human abdominal adipose-derived stem cells (hADSCs) in vitro. A general matrix metalloproteinase (MMP) inhibitor or a MMP2 inhibitor significantly inhibited the PDGF-induced migration. PDGF treatment exhibited greater mRNA level and denser protein level of MMP1. The conditioned medium of PDGF-treated cells showed a caseinolytic activity of MMP1. Transfection of cells with siRNA against MMP1 significantly inhibited MMP1 expression, its caseinolytic activity, and cell migration following PDGF treatment. Phosphatidylinositol 3-kinase (PI3K) inhibitor reduced the migration by about 50% without affecting ERK and MLC proteins. Rho-associated protein kinase inhibitor mostly abolished the migration and MLC proteins. The results suggest that PDGF might signal hADSCs through PI3K, and MMP1 activity could play an important role in this PDGF-induced migration in vitro.

  13. Involvement of the PI3K and ERK signaling pathways in largemouth bass virus-induced apoptosis and viral replication.

    PubMed

    Huang, Xiaohong; Wang, Wei; Huang, Youhua; Xu, Liwen; Qin, Qiwei

    2014-12-01

    Increased reports demonstrated that largemouth Bass, Micropterus salmoides in natural and artificial environments were always suffered from an emerging iridovirus disease, largemouth Bass virus (LMBV). However, the underlying mechanism of LMBV pathogenesis remained largely unknown. Here, we investigated the cell signaling events involved in virus induced cell death and viral replication in vitro. We found that LMBV infection in epithelioma papulosum cyprini (EPC) cells induced typical apoptosis, evidenced by the appearance of apoptotic bodies, cytochrome c release, mitochondrial membrane permeabilization (MMP) destruction and reactive oxygen species (ROS) generation. Two initiators of apoptosis, caspase-8 and caspase-9, and the executioner of apoptosis, caspase-3, were all significantly activated with the infection time, suggested that not only mitochondrion-mediated, but also death receptor-mediated apoptosis were involved in LMBV infection. Reporter gene assay showed that the promoter activity of transcription factors including p53, NF-κB, AP-1 and cAMP response element-binding protein (CREB) were decreased during LMBV infection. After treatment with different signaling pathway inhibitors, virus production were significantly suppressed by the inhibition of phosphatidylinositol 3-kinase (PI3K) pathway and extracellular-signal-regulated kinases (ERK) signaling pathway. Furthermore, LMBV infection induced apoptosis was enhanced by PI3K inhibitor LY294002, but decreased by addition of ERK inhibitor UO126. Therefore, we speculated that apoptosis was sophisticatedly regulated by a series of cell signaling events for efficient virus propagation. Taken together, our results provided new insights into the molecular mechanism of ranavirus infection.

  14. Disruption of PDGFRalpha-initiated PI3K activation and migration of somite derivatives leads to spina bifida.

    PubMed

    Pickett, Elizabeth A; Olsen, Gregory S; Tallquist, Michelle D

    2008-02-01

    Spina bifida, or failure of the vertebrae to close at the midline, is a common congenital malformation in humans that is often synonymous with neural tube defects (NTDs). However, it is likely that other etiologies exist. Genetic disruption of platelet-derived growth factor receptor (PDGFR) alpha results in spina bifida, but the underlying mechanism has not been identified. To elucidate the cause of this birth defect in PDGFRalpha mutant embryos, we examined the developmental processes involved in vertebrae formation. Exposure of chick embryos to the PDGFR inhibitor imatinib mesylate resulted in spina bifida in the absence of NTDs. We next examined embryos with a tissue-specific deletion of the receptor. We found that loss of the receptor from chondrocytes did not recapitulate the spina bifida phenotype. By contrast, loss of the receptor from all sclerotome and dermatome derivatives or disruption of PDGFRalpha-driven phosphatidyl-inositol 3' kinase (PI3K) activity resulted in spina bifida. Furthermore, we identified a migration defect in the sclerotome as the cause of the abnormal vertebral development. We found that primary cells from these mice exhibited defects in PAK1 activation and paxillin localization. Taken together, these results indicate that PDGFRalpha downstream effectors, especially PI3K, are essential for cell migration of a somite-derived dorsal mesenchyme and disruption of receptor signaling in these cells leads to spina bifida.

  15. Dehydroepiandrosterone-induced proliferation of prostatic epithelial cell is mediated by NFKB via PI3K/AKT signaling pathway.

    PubMed

    Sun, Hong-Zhi; Yang, Tong-Wei; Zang, Wei-Jin; Wu, Shu-Fang

    2010-03-01

    Dehydroepiandrosterone (DHEA) is an endogenous steroid that is metabolized to androgens and/or estrogens in the human prostate. DHEA levels decline with age, and use of DHEA supplements to retard the aging process is of unproved effectiveness and safety. In this study, rat ventral prostatic epithelial cells were used to determine whether DHEA-modulated proliferation and prostate-specific antigen (PSA listed as KLKB1 in the MGI Database) production were mediated via the androgen receptor (AR) and its potential mechanism. We demonstrated that proliferation of prostatic epithelial cells and increase of PSA expression induced by DHEA were neutralized by Casodex or Ar siRNA, two specific AR blockers. DHEA stimulated Nfkb DNA binding activity, with this effect being blunted by Casodex or Ar siRNA. Moreover, the inhibition of the phosphatidylinositol 3-kinase (PI3K)/AKT nullified the effects of DHEA on NFKB activation. These findings suggested that DHEA stimulated normal prostatic epithelial cell proliferation, and AR is involved in DHEA-induced PSA expression in normal prostatic epithelial cells. This stimulation effect induced by DHEA is mediated by the activation of NFKB via PI3K/AKT pathway.

  16. Wortmannin potentiates roscovitine-induced growth inhibition in human solid tumor cells by repressing PI3K/Akt pathway.

    PubMed

    Zhang, Feng; Zhang, Tao; Jiang, Tao; Zhang, Rong; Teng, Zeng-hui; Li, Chen; Gu, Zhong-Ping; Mei, Qibing

    2009-12-28

    Roscovitine has been reported to have anti-tumor effects in some cancer cell lines. The phosphatidylinositol-3-kinase (PI3K) signaling, which activates protein kinase B (PKB)/Akt, is known to mediate cell survival. The current study examined the role of wortmannin, a PI3K inhibitor, as a chemosensitizer for roscovitine and its proposed mechanism of action. The results showed that wortmannin significantly chemosensitized three human tumor cell lines (A549, HCT116 and HeLa cells). In A549 cells, wortmannin increased roscovitine-induced apoptosis in a dose-dependent manner, which was correlated with the inhibition of phosphorylated PKB/Akt level. Wortmannin enhanced the effects of roscovitine by causing pronounced reduction of mitochondrial transmembrane potential (MMP) and increases of cytochrome c release and active caspase-3, as well as enhanced activation of Bax and Bad, including Bax oligomerization and mitochondrial translocation of Bax and Bad. Taken together, these results provide evidence for the potential application of roscovitine/wormannin combination in clinical treatment for solid tumors.

  17. PARK2 Depletion Connects Energy and Oxidative Stress to PI3K/Akt Activation via PTEN S-Nitrosylation.

    PubMed

    Gupta, Amit; Anjomani-Virmouni, Sara; Koundouros, Nikos; Dimitriadi, Maria; Choo-Wing, Rayman; Valle, Adamo; Zheng, Yuxiang; Chiu, Yu-Hsin; Agnihotri, Sameer; Zadeh, Gelareh; Asara, John M; Anastasiou, Dimitrios; Arends, Mark J; Cantley, Lewis C; Poulogiannis, George

    2017-03-16

    PARK2 is a gene implicated in disease states with opposing responses in cell fate determination, yet its contribution in pro-survival signaling is largely unknown. Here we show that PARK2 is altered in over a third of all human cancers, and its depletion results in enhanced phosphatidylinositol 3-kinase/Akt (PI3K/Akt) activation and increased vulnerability to PI3K/Akt/mTOR inhibitors. PARK2 depletion contributes to AMPK-mediated activation of endothelial nitric oxide synthase (eNOS), enhanced levels of reactive oxygen species, and a concomitant increase in oxidized nitric oxide levels, thereby promoting the inhibition of PTEN by S-nitrosylation and ubiquitination. Notably, AMPK activation alone is sufficient to induce PTEN S-nitrosylation in the absence of PARK2 depletion. Park2 loss and Pten loss also display striking cooperativity to promote tumorigenesis in vivo. Together, our findings reveal an important missing mechanism that might account for PTEN suppression in PARK2-deficient tumors, and they highlight the importance of PTEN S-nitrosylation in supporting cell survival and proliferation under conditions of energy deprivation. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Sensitivity to PI3K and AKT inhibitors is mediated by divergent molecular mechanisms in subtypes of DLBCL.

    PubMed

    Erdmann, Tabea; Klener, Pavel; Lynch, James T; Grau, Michael; Vočková, Petra; Molinsky, Jan; Tuskova, Diana; Hudson, Kevin; Polanska, Urszula M; Grondine, Michael; Mayo, Michele; Dai, Beiying; Pfeifer, Matthias; Erdmann, Kristian; Schwammbach, Daniela; Zapukhlyak, Myroslav; Staiger, Annette M; Ott, German; Berdel, Wolfgang E; Davies, Barry R; Cruzalegui, Francisco; Trneny, Marek; Lenz, Peter; Barry, Simon T; Lenz, Georg

    2017-07-20

    Activated B-cell-like (ABC) and germinal center B-cell-like diffuse large B-cell lymphoma (DLBCL) represent the 2 major molecular DLBCL subtypes. They are characterized by differences in clinical course and by divergent addiction to oncogenic pathways. To determine activity of novel compounds in these 2 subtypes, we conducted an unbiased pharmacologic in vitro screen. The phosphatidylinositol-3-kinase (PI3K) α/δ (PI3Kα/δ) inhibitor AZD8835 showed marked potency in ABC DLBCL models, whereas the protein kinase B (AKT) inhibitor AZD5363 induced apoptosis in PTEN-deficient DLBCLs irrespective of their molecular subtype. These in vitro results were confirmed in various cell line xenograft and patient-derived xenograft mouse models in vivo. Treatment with AZD8835 induced inhibition of nuclear factor κB signaling, prompting us to combine AZD8835 with the Bruton's tyrosine kinase inhibitor ibrutinib. This combination was synergistic and effective both in vitro and in vivo. In contrast, the AKT inhibitor AZD5363 was effective in PTEN-deficient DLBCLs through downregulation of the oncogenic transcription factor MYC. Collectively, our data suggest that patients should be stratified according to their oncogenic dependencies when treated with PI3K and AKT inhibitors. © 2017 by The American Society of Hematology.

  19. Neurotrophin Promotes Neurite Outgrowth by Inhibiting Rif GTPase Activation Downstream of MAPKs and PI3K Signaling

    PubMed Central

    Tian, Xiaoxia; Yan, Huijuan; Li, Jiayi; Wu, Shuang; Wang, Junyu; Fan, Lifei

    2017-01-01

    Members of the well-known semaphorin family of proteins can induce both repulsive and attractive signaling in neural network formation and their cytoskeletal effects are mediated in part by small guanosine 5’-triphosphatase (GTPases). The aim of this study was to investigate the cellular role of Rif GTPase in the neurotrophin-induced neurite outgrowth. By using PC12 cells which are known to cease dividing and begin to show neurite outgrowth responding to nerve growth factor (NGF), we found that semaphorin 6A was as effective as nerve growth factor at stimulating neurite outgrowth in PC12 cells, and that its neurotrophic effect was transmitted through signaling by mitogen-activated protein kinases (MAPKs) and phosphatidylinositol-3-kinase (PI3K). We further found that neurotrophin-induced neurite formation in PC12 cells could be partially mediated by inhibition of Rif GTPase activity downstream of MAPKs and PI3K signaling. In conclusion, we newly identified Rif as a regulator of the cytoskeletal rearrangement mediated by semaphorins. PMID:28098758

  20. Neurotrophin Promotes Neurite Outgrowth by Inhibiting Rif GTPase Activation Downstream of MAPKs and PI3K Signaling.

    PubMed

    Tian, Xiaoxia; Yan, Huijuan; Li, Jiayi; Wu, Shuang; Wang, Junyu; Fan, Lifei

    2017-01-13

    Members of the well-known semaphorin family of proteins can induce both repulsive and attractive signaling in neural network formation and their cytoskeletal effects are mediated in part by small guanosine 5'-triphosphatase (GTPases). The aim of this study was to investigate the cellular role of Rif GTPase in the neurotrophin-induced neurite outgrowth. By using PC12 cells which are known to cease dividing and begin to show neurite outgrowth responding to nerve growth factor (NGF), we found that semaphorin 6A was as effective as nerve growth factor at stimulating neurite outgrowth in PC12 cells, and that its neurotrophic effect was transmitted through signaling by mitogen-activated protein kinases (MAPKs) and phosphatidylinositol-3-kinase (PI3K). We further found that neurotrophin-induced neurite formation in PC12 cells could be partially mediated by inhibition of Rif GTPase activity downstream of MAPKs and PI3K signaling. In conclusion, we newly identified Rif as a regulator of the cytoskeletal rearrangement mediated by semaphorins.

  1. Endomembrane PtdIns(3,4,5)P3 activates the PI3K-Akt pathway.

    PubMed

    Jethwa, Nirmal; Chung, Gary H C; Lete, Marta G; Alonso, Alicia; Byrne, Richard D; Calleja, Véronique; Larijani, Banafshé

    2015-09-15

    PKB/Akt activation is a common step in tumour growth, proliferation and survival. Akt activation is understood to occur at the plasma membrane of cells in response to growth factor stimulation and local production of the phosphoinositide lipid phosphatidylinositol (3,4,5)-trisphosphate [PtdIns(3,4,5)P3] following phosphoinositide 3-kinase (PI3K) activation. The metabolism and turnover of phosphoinositides is complex--they act as signalling molecules as well as structural components of biological membranes. The localisation and significance of internal pools of PtdIns(3,4,5)P3 has long been speculated upon. By using transfected and recombinant protein probes for PtdIns(3,4,5)P3, we show that PtdIns(3,4,5)P3 is enriched in the nuclear envelope and early endosomes. By exploiting an inducible dimerisation device to recruit Akt to these compartments, we demonstrate that Akt can be locally activated in a PtdIns(3,4,5)P3-dependent manner and has the potential to phosphorylate compartmentally localised downstream substrates. This could be an important mechanism to regulate Akt isoform substrate specificity or influence the timing and duration of PI3K pathway signalling. Defects in phosphoinositide metabolism and localisation are known to contribute to cancer, suggesting that interactions at subcellular compartments might be worthwhile targets for therapeutic intervention.

  2. PI3K regulates endocytosis after insulin secretion by mediating signaling crosstalk between Arf6 and Rab27a.

    PubMed

    Yamaoka, Mami; Ando, Tomomi; Terabayashi, Takeshi; Okamoto, Mitsuhiro; Takei, Masahiro; Nishioka, Tomoki; Kaibuchi, Kozo; Matsunaga, Kohichi; Ishizaki, Ray; Izumi, Tetsuro; Niki, Ichiro; Ishizaki, Toshimasa; Kimura, Toshihide

    2016-02-01

    In secretory cells, endocytosis is coupled to exocytosis to enable proper secretion. Although endocytosis is crucial to maintain cellular homeostasis before and after secretion, knowledge about secretagogue-induced endocytosis in secretory cells is still limited. Here, we searched for proteins that interacted with the Rab27a GTPase-activating protein (GAP) EPI64 (also known as TBC1D10A) and identified the Arf6 guanine-nucleotide-exchange factor (GEF) ARNO (also known as CYTH2) in pancreatic β-cells. We found that the insulin secretagogue glucose promotes phosphatidylinositol (3,4,5)-trisphosphate (PIP3) generation through phosphoinositide 3-kinase (PI3K), thereby recruiting ARNO to the intracellular side of the plasma membrane. Peripheral ARNO promotes clathrin assembly through its GEF activity for Arf6 and regulates the early stage of endocytosis. We also found that peripheral ARNO recruits EPI64 to the same area and that the interaction requires glucose-induced endocytosis in pancreatic β-cells. Given that GTP- and GDP-bound Rab27a regulate exocytosis and the late stage of endocytosis, our results indicate that the glucose-induced activation of PI3K plays a pivotal role in exocytosis-endocytosis coupling, and that ARNO and EPI64 regulate endocytosis at distinct stages.

  3. Salidroside attenuates colistin-induced neurotoxicity in RSC96 Schwann cells through PI3K/Akt pathway.

    PubMed

    Lu, Ziyin; Jiang, Guozheng; Chen, Ying; Wang, Jian; Muhammad, Ishfaq; Zhang, Ling; Wang, Rui; Liu, Fangping; Li, Rui; Qian, Feng; Li, Jichang

    2017-06-01

    Neurotoxicity is a key dose-limiting factor for colistin therapy. This study aimed to investigate the protective effect of Salidroside on colistin-induced neurotoxicity in RSC96 Schwann cells and the underlying mechanisms. After Salidroside (12.5, 25, 50 μg/mL) treatment for 2 h, the cells were cultured with 250 μg/mL colistin for 24 h. In order to investigate the role of phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway, the cells were pre-treated with LY294002 (12.5 μmol/L, a specific inhibitor of PI3K phosphorylation) for 1 h before Salidroside (50 μg/mL) treatment, then were co-cultured with colistin (250 μg/mL) for 24 h. The results showed that colistin treatment could induce apoptotic cell death which was associated with oxidative stress injury. Salidroside could reduce colistin-induced neurotoxicity, decrease the effect of colistin on the reduced expression levels of p-Akt and Bcl-2, and increased the expresion of Bax, release of Cyt c, and activation of caspase-3. However, the protective effect of Salidroside against colistin-induced apoptosis was partly abolished by LY294002. These findings suggest that Salidroside could attenuate colistin-induced neurotoxicity in RSC96 Schwann cells via the PI3K/Akt pathway. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Garlic Oil Suppressed Nitrosodiethylamine-Induced Hepatocarcinoma in Rats by Inhibiting PI3K-AKT-NF-κB Pathway.

    PubMed

    Zhang, Cui-Li; Zeng, Tao; Zhao, Xiu-Lan; Xie, Ke-Qin

    2015-01-01

    To explore the underlying mechanisms for the protective effects of garlic oil (GO) against nitrosodiethylamine (NDEA)-induced hepatocarcinoma, 60 male Wistar rats were randomized into 4 groups (n=15): control group, NDEA group, and two GO plus NDEA groups. The rats in GO plus NDEA groups were pretreated with GO (20 or 40 mg/kg) for 7 days. Then, all rats except those in control group were gavaged with NDEA for 20 weeks, and the rats in GO plus NDEA groups were continuously administered with GO. The results showed that GO co-treatment significantly suppressed the NDEA-induced increases of alpha fetal protein (AFP) level in serum, nuclear atypia in H&E staining, sirius red-positive areas and proliferating cell nuclear antigen (PCNA) expression. The molecular mechanisms exploration revealed that the protein levels of phosphatidylinositol 3 kinase (PI3K)-p85, PI3K-p110, total AKT, p-AKT (Ser473) and p-AKT (Thr308) in the liver of NDEA group rats were higher than those in control group rats. In addition, NDEA treatment induced IκB degradation and NF-κB p65 phosphorylation, and up-regulated the protein levels of downstream pro-inflammatory mediators. GO co-treatment significantly reversed all the above adverse effects induced by NDEA. These results suggested that the protective effects of GO against NDEA-induced hepatocarcinoma might be associated with the suppression of PI3K- AKT-NF-κB pathway.

  5. Effects of dihydrotestosterone on adhesion and proliferation via PI3-K/Akt signaling in endothelial progenitor cells.

    PubMed

    Liu, Rui; Ding, Li; Yu, Ming-Hua; Wang, Han-Qin; Li, Wen-Chun; Cao, Zheng; Zhang, Peng; Yao, Bo-Chun; Tang, Jie; Ke, Qing; Huang, Tie-Zhu

    2014-08-01

    The protective effects of male hormones on the cardiovascular system are still in dispute. There is now ample evidence that testosterone level is negatively correlated to the incidence and mortality of cardiovascular disease in men. Endothelial progenitor cells (EPCs) play a vital role in endothelial healing and vascular integrity, which are useful for promoting cardiovascular health. In this study, we investigated the effects of dihydrotestosterone (DHT), a non-aromatizable androgen, on human EPC function and the activation of the phosphatidylinositol-3-kinase (PI3-K)/Akt pathway in vitro. EPCs were incubated with a series of concentrations (1, 10, or 100 nmol/L in DMSO) of DHT for 24 h or with 10 nmol/L DHT for different time (6, 12, 24, 48 h). EPC adhesion and proliferation and the activation of Akt were assayed by cell counting, 5-ethynyl-2'-deoxyuridine incorporation assay, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and Western blot analysis. Our data demonstrated that DHT significantly increased the proliferative activity and adhesive ability of EPCs in a dose- and time-dependent manner, maximum at 10 nmol/L, 24 h (p < 0.05). Western blot analysis revealed that DHT promoted the phosphorylation of Akt, and the effects of different concentrations of DHT on Akt phosphorylation were consistent with those on EPC proliferation and adhesion (p < 0.05). However, the enhancing effects of DHT on EPCs decreased with administration of the pharmacological PI3-K blocker LY294002 (p < 0.05). In conclusion, DHT can modulate EPC proliferation and adhesion and the PI3-K/Akt pathway plays an important role in this process.

  6. Garlic Oil Suppressed Nitrosodiethylamine-Induced Hepatocarcinoma in Rats by Inhibiting PI3K-AKT-NF-κB Pathway

    PubMed Central

    Zhang, Cui-Li; Zeng, Tao; Zhao, Xiu-Lan; Xie, Ke-Qin

    2015-01-01

    To explore the underlying mechanisms for the protective effects of garlic oil (GO) against nitrosodiethylamine (NDEA)-induced hepatocarcinoma, 60 male Wistar rats were randomized into 4 groups (n=15): control group, NDEA group, and two GO plus NDEA groups. The rats in GO plus NDEA groups were pretreated with GO (20 or 40 mg/kg) for 7 days. Then, all rats except those in control group were gavaged with NDEA for 20 weeks, and the rats in GO plus NDEA groups were continuously administered with GO. The results showed that GO co-treatment significantly suppressed the NDEA-induced increases of alpha fetal protein (AFP) level in serum, nuclear atypia in H&E staining, sirius red-positive areas and proliferating cell nuclear antigen (PCNA) expression. The molecular mechanisms exploration revealed that the protein levels of phosphatidylinositol 3 kinase (PI3K)-p85, PI3K-p110, total AKT, p-AKT (Ser473) and p-AKT (Thr308) in the liver of NDEA group rats were higher than those in control group rats. In addition, NDEA treatment induced IκB degradation and NF-κB p65 phosphorylation, and up-regulated the protein levels of downstream pro-inflammatory mediators. GO co-treatment significantly reversed all the above adverse effects induced by NDEA. These results suggested that the protective effects of GO against NDEA-induced hepatocarcinoma might be associated with the suppression of PI3K- AKT-NF-κB pathway. PMID:25999787

  7. Targeting EMP3 suppresses proliferation and invasion of hepatocellular carcinoma cells through inactivation of PI3K/Akt pathway

    PubMed Central

    Lee, Chien-Hsing; Yang, Shun-Fa; Cheng, Chun-Wen; Tang, Meng-Ju; Lin, Chia-Liang; Lin, Chu-Liang; Chou, Ruey-Hwang

    2015-01-01

    Epithelial membrane protein-3 (EMP3), a typical member of the epithelial membrane protein (EMP) family, is epigenetically silenced in some cancer types, and has been proposed to be a tumor suppressor gene. However, its effects on tumor suppression are controversial and its roles in development and malignancy of hepatocellular carcinoma (HCC) remain unclear. In the present study, we found that EMP3 was highly expressed in the tumorous tissues comparing to the matched normal tissues, and negatively correlated with differentiated degree of HCC patients. Knockdown of EMP3 significantly reduced cell proliferation, arrested cell cycle at G1 phase, and inhibited the motility and invasiveness in accordance with the decreased expression and activity of urokinase plasminogen activator (uPA) and matrix metalloproteinase 9 (MMP-9) in HCC cells. The in vivo tumor growth of HCC was effectively suppressed by knockdown of EMP3 in a xenograft mouse model. The EMP3 knockdown-reduced cell proliferation and invasion were attenuated by inhibition of phosphatidylinositol 3-kinase (PI3K) or knockdown of Akt, and rescued by overexpression of Akt in HCC cells. Clinical positive correlations of EMP3 with p85 regulatory subunit of PI3K, p-Akt, uPA, as well as MMP-9 were observed in the tissue sections from HCC patients. Here, we elucidated the tumor progressive effects of EMP3 through PI3K/Akt pathway and uPA/MMP-9 cascade in HCC cells. The findings provided a new insight into EMP3, which might be a potential molecular target for diagnosis and treatment of HCC. PMID:26472188

  8. Activation of the PI3K/mTOR/AKT Pathway and Survival in Solid Tumors: Systematic Review and Meta-Analysis

    PubMed Central

    Ocana, Alberto; Vera-Badillo, Francisco; Al-Mubarak, Mustafa; Templeton, Arnoud J.; Corrales-Sanchez, Verónica; Diez-Gonzalez, Laura; Cuenca-Lopez, María D.; Seruga, Bostjan; Pandiella, Atanasio; Amir, Eitan

    2014-01-01

    Background Aberrations in the phosphatidylinositol 3-kinase (PI3K)/mammalian target of rapamycin (mTOR)/AKT pathway are common in solid tumors. Numerous drugs have been developed to target different components of this pathway. However the prognostic value of these aberrations is unclear. Methods PubMed was searched for studies evaluating the association between activation of the PI3K/mTOR/AKT pathway (defined as PI3K mutation [PIK3CA], lack of phosphatase and tensin homolog [PTEN] expression by immunohistochemistry or western-blot or increased expression/activation of downstream components of the pathway by immunohistochemistry) with overall survival (OS) in solid tumors. Published data were extracted and computed into odds ratios (OR) for death at 5 years. Data were pooled using the Mantel-Haenszel random-effect model. Results Analysis included 17 studies. Activation of the PI3K/mTOR/AKT pathway was associated with significantly worse 5-year survival (OR:2.12, 95% confidence intervals 1.42–3.16, p<0.001). Loss of PTEN expression and increased expression/activation of downstream components were associated with worse survival. No association between PIK3CA mutations and survival was observed. Differences between methods for assessing activation of the PI3K/mTOR/AKT pathway were statistically significant (p = 0.04). There was no difference in the effect of up-regulation of the pathway on survival between different cancer sites (p = 0.13). Conclusion Activation of the PI3K/AKT/mTOR pathway, especially if measured by loss of PTEN expression or increased expression/activation of downstream components is associated with poor survival. PIK3CA mutational status is not associated with adverse outcome, challenging its value as a biomarker of patient outcome or as a stratification factor for patients treated with agents acting on the PI3K/AKT/mTOR pathway. PMID:24777052

  9. Jaceosidin, a natural flavone, promotes angiogenesis via activation of VEGFR2/FAK/PI3K/AKT/NF-κB signaling pathways in endothelial cells.

    PubMed

    Lee, Tae Hoon; Jung, Hana; Park, Keun Hyung; Bang, Myun Ho; Baek, Nam-In; Kim, Jiyoung

    2014-10-01

    Angiogenesis, the growth of new blood vessels from pre-existing vasculature, plays an important role in physiological and pathological processes such as embryonic development wound healing and revascularization of tissues after exposure to ischemia. We investigated the effects of jaceosidin, a main constituent of medicinal herbs of the genus Artemisia, on angiogenesis and signaling pathways in endothelial cells. Jaceosidin stimulated proliferation, migration and tubulogenesis of ECs as well as ex vivo sprouting from aorta rings, which are phenomena typical of angiogenesis. Jaceosidin activated vascular endothelial growth factor receptor 2 (VEGFR2, FLk-1/KDR) and angiogenic signaling molecules such as focal adhesion kinase, phosphatidylinositol 3-kinase, and its downstream target, the serine-threonine kinase AKTWe also demonstrated that jaceosidin activated the NF-κB-driven expression of a luciferase reporter gene and NF-κB binding to DNA. Jaceosidin-induced proliferation and migration of human umbilical vascular endothelial cells were strongly inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002 and NF-κB inhibitor BAY11-7082, indicating that the PI3K/AKT/NF-κB signaling pathway is involved in jaceosidin-induced angiogenesis. Our results suggest that jaceosidin stimulates angiogenesis by activating the VEGFR2/FAK/PI3K/AKT/NF-κB signaling pathway and that it may be useful in developing angiogenic agents to promote the growth of collateral blood vessels in ischemic tissues.

  10. [6]-Shogaol Inhibits α-MSH-Induced Melanogenesis through the Acceleration of ERK and PI3K/Akt-Mediated MITF Degradation

    PubMed Central

    Huang, Huey-Chun; Chang, Shu-Jen; Wu, Chia-Yin; Ke, Hui-Ju; Chang, Tsong-Min

    2014-01-01

    [6]-Shogaol is the main biologically active component of ginger. Previous reports showed that [6]-shogaol has several pharmacological characteristics, such as antioxidative, anti-inflammatory, antimicrobial, and anticarcinogenic properties. However, the effects of [6]-shogaol on melanogenesis remain to be elucidated. The study aimed to evaluate the potential skin whitening mechanisms of [6]-shogaol. The effects of [6]-shogaol on cell viability, melanin content, tyrosinase activity, and the expression of the tyrosinase and microphthalmia-associated transcription factor (MITF) were measured. The results revealed that [6]-shogaol effectively suppresses tyrosinase activity and the amount of melanin and that those effects are more pronounced than those of arbutin. It was also found that [6]-shogaol decreased the protein expression levels of tyrosinase-related protein 1 (TRP-1) and microphthalmia-associated transcriptional factor (MITF). In addition, the MITF mRNA levels were also effectively decreased in the presence of 20 μM [6]-shogaol. The degradation of MITF protein was inhibited by the MEK 1-inhibitor (U0126) or phosphatidylinositol-3-kinase inhibitor (PI3K inhibitor) (LY294002). Further immunofluorescence staining assay implied the involvement of the proteasome in the downregulation of MITF by [6]-shogaol. Our confocal assay results also confirmed that [6]-shogaol inhibited α-melanocyte stimulating hormone- (α-MSH-) induced melanogenesis through the acceleration of extracellular responsive kinase (ERK) and phosphatidylinositol-3-kinase- (PI3K/Akt-) mediated MITF degradation. PMID:25045707

  11. Oleanolic acid supplement attenuates liquid fructose-induced adipose tissue insulin resistance through the insulin receptor substrate-1/phosphatidylinositol 3-kinase/Akt signaling pathway in rats

    SciTech Connect

    Li, Ying; Wang, Jianwei; Gu, Tieguang; Yamahara, Johji; Li, Yuhao

    2014-06-01

    fructose-fed rats. • OA attenuated fructose-induced increase in Adipo-IR index and NEFA concentrations. • OA modulated adipose IRS-1/phosphatidylinositol 3-kinase/Akt signaling. • OA ameliorates Adipo-IR via the IRS-1/PI3K/Akt signaling pathway in rats.

  12. Structure of the iSH2 domain of Human phosphatidylinositol 3-kinase p85 beta Subunit Reveals Conformational Plasticity in the Interhelical Turn Region

    SciTech Connect

    C Schauder; L Ma; R Krug; G Montelione; R Guan

    2011-12-31

    Phosphatidylinositol 3-kinase (PI3K) proteins actively trigger signaling pathways leading to cell growth, proliferation and survival. These proteins have multiple isoforms and consist of a catalytic p110 subunit and a regulatory p85 subunit. The iSH2 domain of the p85 {beta} isoform has been implicated in the binding of nonstructural protein 1 (NS1) of influenza A viruses. Here, the crystal structure of human p85 {beta} iSH2 determined to 3.3 {angstrom} resolution is reported. The structure reveals that this domain mainly consists of a coiled-coil motif. Comparison with the published structure of the bovine p85 {beta} iSH2 domain bound to the influenza A virus nonstructural protein 1 indicates that little or no structural change occurs upon complex formation. By comparing this human p85 {beta} iSH2 structure with the bovine p85 {beta} iSH2 domain, which shares 99% sequence identity, and by comparing the multiple conformations observed within the asymmetric unit of the bovine iSH2 structure, it was found that this coiled-coil domain exhibits a certain degree of conformational variability or 'plasticity' in the interhelical turn region. It is speculated that this plasticity of p85 {beta} iSH2 may play a role in regulating its functional and molecular-recognition properties.

  13. Polycystin-1 Induces Cell Migration by Regulating Phosphatidylinositol 3-kinase-dependent Cytoskeletal Rearrangements and GSK3β-dependent Cell–Cell Mechanical Adhesion

    PubMed Central

    Boca, Manila; D'Amato, Lisa; Distefano, Gianfranco; Polishchuk, Roman S.; Germino, Gregory G.

    2007-01-01

    Polycystin-1 (PC-1) is a large plasma-membrane receptor encoded by the PKD1 gene mutated in autosomal dominant polycystic kidney disease (ADPKD). Although the disease is thought to be recessive on a molecular level, the precise mechanism of cystogenesis is unclear, although cytoarchitecture defects seem to be the most likely initiating events. Here we show that PC-1 regulates the actin cytoskeleton in renal epithelial cells (MDCK) and induces cell scattering and cell migration. All of these effects require phosphatidylinositol 3-kinase (PI3-K) activity. Consistent with these observations Pkd1−/− mouse embryonic fibroblasts (MEFs) have reduced capabilities to migrate compared with controls. PC-1 overexpressing MDCK cells are able to polarize normally with proper adherens and tight junctions formation, but show quick reabsorption of ZO-1, E-cadherin, and β-catenin upon wounding of a monolayer and a transient epithelial-to-mesenchymal transition (EMT) that favors a rapid closure of the wound and repolarization. Finally, we show that PC-1 is able to control the turnover of cytoskeletal-associated β-catenin through activation of GSK3β. Expression of a nondegradable form of β-catenin in PC-1 MDCK cells restores strong cell–cell mechanical adhesion. We propose that PC-1 might be a central regulator of epithelial plasticity and its loss results in impaired normal epithelial homeostasis. PMID:17671167

  14. PI3K/AKT inhibition induces caspase-dependent apoptosis in HTLV-1-transformed cells.

    PubMed

    Jeong, Soo-Jin; Dasgupta, Arindam; Jung, Kyung-Jin; Um, Jee-Hyun; Burke, Aileen; Park, Hyeon Ung; Brady, John N

    2008-01-20

    The phosphatidylinositol-3-kinase (PI3K) and AKT (protein kinase B) signaling pathways play an important role in regulating cell cycle progression and cell survival. In previous studies, we demonstrated that AKT is activated in HTLV-1-transformed cells and that Tax activation of AKT is linked to p53 inhibition and cell survival. In the present study, we extend these observations to identify regulatory pathways affected by AKT in HTLV-1-transformed cells. We demonstrate that inhibition of AKT reduces the level of phosphorylated Bad, an important member of the pro-apoptotic family of proteins. Consistent with the decrease of phosphorylated Bad, cytochrome c is released from the mitochondria and caspase-9 is activated. Pretreatment of the cells with caspase-9 specific inhibitor z-LEHD-FMK or pan caspase inhibitor Ac-DEVD-CHO prevented LY294002-induced apoptosis. Of interest, p53 siRNA prevents LY294002-induced apoptosis in HTLV-1-transformed cells, suggesting that p53 reactivation is linked to apoptosis. In conclusion, the AKT pathway is involved in targeting multiple proteins which regulate caspase- and p53-dependent apoptosis in HTLV-1-transformed cells. Since AKT inhibitors simultaneously inhibit NF-kappaB and activate p53, these drugs should be promising candidates for HTLV-1-associated cancer therapy.

  15. Naringin inhibits TNF-α induced oxidative stress and inflammatory response in HUVECs via Nox4/NF-κ B and PI3K/Akt pathways.

    PubMed

    Li, Wenshuang; Wang, Changyuan; Peng, Jinyong; Liang, Jing; Jin, Yue; Liu, Qi; Meng, Qiang; Liu, Kexin; Sun, Huijun

    2014-01-01

    In the development of atherosclerosis, naringin has exhibited potential protective effects. However, the specific mechanisms are not clearly understood. The aim of this trial was to determine the anti-oxidative and anti-inflammatory effects of naringin and uncover the mechanisms in Tumor Necrosis Factor-alpha (TNF-α) induced Human Umbilical Vein Endothelial Cells (HUVECs). Reactive Oxygen Species (ROS) were measured by flow cytometry assay. The levels of NADPH oxidase 4 (Nox4), p22(phox), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) over-expressions were measured by qRT-PCR and Western blotting analyses. Activation of Phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and Nuclear Factor-κB (NF-κB) was evaluated by Western blotting. Naringin inhibited ROS production as well as over-expression levels of Nox4, p22(phox) induced by TNF-α. Naringin inhibited TNF-α induced mRNA and protein over-expressions of ICAM-1 and VCAM-1. Naringin also suppressed activation of NF-κB and PI3K/Akt signaling pathways. These results indicated the preventive effects of naringin on HUVECs injury caused by oxidative stress and inflammation response and the effects might be obtained via inhibition of Nox4 and NF-κB pathways as well as activation of PI3K/Akt pathway. Naringin may be useful in preventing endothelial dysfunction, therefore to ameliorate the development of atherosclerosis.

  16. Combined effects of PI3K and SRC kinase inhibitors with imatinib on intracellular calcium levels, autophagy, and apoptosis in CML-PBL cells.

    PubMed

    Ciarcia, Roberto; Damiano, Sara; Montagnaro, Serena; Pagnini, Ugo; Ruocco, Antonio; Caparrotti, Giuseppe; d'Angelo, Danila; Boffo, Silvia; Morales, Fátima; Rizzolio, Flavio; Florio, Salvatore; Giordano, Antonio

    2013-09-01

    Imatinib induces a complete cytogenetic regression in a large percentage of patients affected by chronic myeloid leukemia (CML) until mutations in the kinase domain of BCR-ABL appear. Alternative strategies for CML patients include the inhibition of phosphatidylinositol 3-kinase (PI3K)-Akt-mammalian target of rapamycin (mTOR) pathway, which is constitutively activated in leukemia cells and seems important for the regulation of cell proliferation, viability, and autophagy. In this study, we verified the effect of imatinib mesylate (IM), alone or in association with LY294002 (LY) (a specific PI3K protein tyrosine kinase inhibitor) or 4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo[3,4-d]-pyrimidine (PP1) (a Src tyrosine kinase inhibitor), on viability, intracellular calcium mobilization, apoptosis, and autophagy, in order to verify possible mechanisms of interaction. Our data demonstrated that PP1 and LY interact synergistically with IM by inducing apoptosis and autophagy in Bcr/Abl+ leukemia cells and this mechanism is related to the stress of the endoplasmic reticulum (ER). Our findings suggest a reasonable relationship between apoptotic and autophagic activity of tyrosine kinase inhibitors (TKIs) and the functionality of smooth ER Ca (2+)-ATPase and inositol triphosphate receptors, independently of intracellular calcium levels. Therapeutic strategies combining imatinib with PI3K and/or Src kinase inhibitors warrant further investigations in Bcr/Abl+ malignancies, particularly in the cases of imatinib mesylate-resistant disease.

  17. Geniposide induces the expression of heme oxygenase-1 via PI3K/Nrf2-signaling to enhance the antioxidant capacity in primary hippocampal neurons.

    PubMed

    Yin, Fei; Liu, Jianhui; Zheng, Xuxu; Guo, Lixia; Xiao, He

    2010-01-01

    Oxidative stress in brain is emerging as a potential causal factor in aging and age-related neurodegenerative disorders. A large body of evidence shows that induction of endogenous antioxidative proteins seems to be a reasonable strategy for delaying the progression of cell injury. In this study, geniposide upregulates the expression of heme oxygenase-1 (HO-1) to attenuate the cell apoptosis induced by 3-morpholinosydnonimine hydrochloride (SIN-1) in primary cultured hippocampal neurons. Furthermore, geniposide induces the nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and activation of phosphatidylinositol 3'-kinase (PI3K) in the presence of oxidative stress, and both LY294002 (a specific inhibitor of PI3K) and Zinc protoporphyrin (ZnPP, an inhibitor of HO-1) decrease the cytoprotective action of geniposide in hippocampal neurons. Taken together, the novel cytoprotective mechanism of geniposide to antagonize oxidative stress may be involved in PI3K- and Nrf2-mediated upregulation of the antioxidative enzyme HO-1.

  18. Rapid-acting antidepressant-like effects of acetyl-l-carnitine mediated by PI3K/AKT/BDNF/VGF signaling pathway in mice.

    PubMed

    Wang, W; Lu, Y; Xue, Z; Li, C; Wang, C; Zhao, X; Zhang, J; Wei, X; Chen, X; Cui, W; Wang, Q; Zhou, W

    2015-01-29

    The possible involvement of the PI3K/AKT/BDNF/VGF signaling in rapid-acting antidepressant-like effects of antidepressants has been explored progressively by more studies. However, whether this signaling participates in the antidepressant-like effects of acetyl-l-carnitine (ALC) has not been examined. Herein, we assessed the antidepressant-like effects of ALC using the forced swimming test (FST). Our results demonstrated the dose-effect relationship of acute administration of ALC (5, 25, 50 and 100mg/kg, i.p.) and showed that it dose-dependently decreased the immobility time on FST of mice. In addition, ALC (100 mg/kg, i.p.) also reversed depressive-like behavior and the down-regulation of phosphorylated AKT (pAKT), brain-derived neurotrophic factor (BDNF) and neuropeptide VGF in the hippocampus and prefrontal cortex of mice induced by chronic unpredictable mild stress (CUMS) paradigm. Further, intra-cerebroventricular (i.c.v.) infusions of LY294002 (10 nmol/side), a specific phosphatidylinositol 3-kinase (PI3K) inhibitor, significantly prevented the antidepressant-like effect of ALC (100mg/kg, i.p.). In conclusion, our results demonstrated that ALC exerts rapid-acting antidepressant-like effects that might be mediated by the PI3K/AKT/BDNF/VGF signaling pathway.

  19. Icariin stimulates angiogenesis by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways in human endothelial cells

    SciTech Connect

    Chung, Byung-Hee; Kim, Jong-Dai; Kim, Chun-Ki; Kim, Jung Huan; Won, Moo-Ho; Lee, Han-Soo; Dong, Mi-Sook; Ha, Kwon-Soo; Kwon, Young-Geun; Kim, Young-Myeong

    2008-11-14

    We investigated the molecular effect and signal pathway of icariin, a major flavonoid of Epimedium koreanum Nakai, on angiogenesis. Icariin stimulated in vitro endothelial cell proliferation, migration, and tubulogenesis, which are typical phenomena of angiogenesis, as well as increased in vivo angiogenesis. Icariin activated the angiogenic signal modulators, ERK, phosphatidylinositol 3-kinase (PI3K), Akt, and endothelial nitric oxide synthase (eNOS), and increased NO production, without affecting VEGF expression, indicating that icariin may directly stimulate angiogenesis. Icariin-induced ERK activation and angiogenic events were significantly inhibited by the MEK inhibitor PD98059, without affecting Akt and eNOS phosphorylation. The PI3K inhibitor Wortmannin suppressed icariin-mediated angiogenesis and Akt and eNOS activation without affecting ERK phosphorylation. Moreover, the NOS inhibitor NMA partially reduced the angiogenic activity of icariin. These results suggest that icariin stimulated angiogenesis by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways and may be a useful drug for angiogenic therapy.

  20. Rat white adipocytes activate p85/p110 PI3K and induce PM GLUT4 in response to adrenoceptor agonists or aluminum fluoride.

    PubMed

    Ohsaka, Y; Nomura, Y

    2016-03-01

    Adipocyte responses to adrenergic and ß-adrenoceptor(-AR) (adrenoceptor) regulation are not sufficiently understood, and information helpful for elucidating the adrenoceptor-responsive machinery is insufficient. Here we show by using immunoprecipitated kinase analysis with a phosphatidylinositol 3-kinase (PI3K) p85 antibody that PI3K activation was induced by treatment with 10 or 100 µM norepinephrine (NE) for 15 min or with 10 mM aluminum fluoride (AF, a guanosine triphosphate (GTP)-binding (G) protein activator) for 20 min in white adipocytes (rat epididymal adipocytes) and that treatment with pertussis toxin (PTX, a G-protein inactivator) inhibited PI3K activation induced by the 20-min treatment with AF in the cells. In addition, western blot analysis revealed that glucose transporter 4 (GLUT4) level in the adipocyte plasma membrane (PM) fraction was increased by treatment with 10 µM NE, 100 µM dobutamine (DOB, a ß1-AR agonist), or 0.1 µM CL316243 (CL, a ß3-AR agonist) for 30 min or with 10 mM AF for 20 min. NE or AF treatment triggered 2-deoxyglucose (2-DG) uptake into adipocytes under the above conditions. Our results advance the understanding of responses to adrenoceptor regulation in white adipocytes and provide possible clues for clarifying the machinery involved in adrenergic and ß-AR responses in the cells.

  1. Protective Effect of Minocycline Against Ketamine-Induced Injury in Neural Stem Cell: Involvement of PI3K/Akt and Gsk-3 Beta Pathway

    PubMed Central

    Lu, Yang; Lei, Shan; Wang, Ning; Lu, Pan; Li, Weisong; Zheng, Juan; Giri, Praveen K.; Lu, Haixia; Chen, Xinlin; Zuo, Zhiyi; Liu, Yong; Zhang, Pengbo

    2016-01-01

    It has been suggested that ketamine cause injury during developing brain. Minocycline (MC) could prevent neuronal cell death through the activation of cell survival signals and the inhibition of apoptotic signals in models of neurodegenerative diseases. Here we investigated the protective effect of MC against ketamine-induced injury in neural stem cells (NSCs) from neonatal rat. Ketamine (100 μM/L) significantly inhibited NSC proliferation, promoted their differentiation into astrocytes and suppressed neuronal differentiation of NSCs. Moreover, the apoptotic level was increased following ketamine exposure. MC pretreatment greatly enhanced cell viability, decreased caspase-3-like activity, even reversed the differentiation changes caused by ketamine. To elucidate a possible mechanism of MC’ neuroprotective effect, we investigated the phosphatidylinositol 3-kinase (PI3K) pathway using LY294002, a specific PI3K inhibitor. Immunoblotting revealed that MC enhanced the phosphorylation/activation of Akt and phosphorylation/inactivation of glycogen synthase kinase-3beta (Gsk-3β). Our results suggest that PI3K/Akt and Gsk-3β pathway are involved in the neuroprotective effect of MC. PMID:28066173

  2. Arctigenin, a Natural Lignan Compound, Induces Apoptotic Death of Hepatocellular Carcinoma Cells via Suppression of PI3-K/Akt Signaling.

    PubMed

    Jiang, Xiaoxin; Zeng, Leping; Huang, Jufang; Zhou, Hui; Liu, Yubin

    2015-04-28

    In this study, we explored the cytotoxic effects of arctigenin, a natural lignan compound, on human hepatocellular carcinoma (HCC) cells and check the involvement of phosphatidylinositol 3-kinase (PI3-K)/Akt signaling. HCC cells were treated with different concentrations of arctigenin and cell viability and apoptosis were assessed. Manipulating Akt signaling was used to determine its role in the action of arctigenin. Arctigenin significantly inhibited the viability of HCC cells in a concentration-dependent manner. Arctigenin induced apoptosis and activation of caspase-9 and -3. Overexpression of a constitutively active Akt mutant blocked arctigenin-induced apoptosis. Combinational treatment with arctigenin and the PI3-K inhibitor LY294002 significantly enhanced apoptosis. Arctigenin reduced the expression of Bcl-xL, Mcl-1, and survivin and the phosphorylation of mTOR and S6K, which were significantly reversed by overexpression of constitutively active Akt. This is the first report about the anticancer activity of arctigenin in HCC cells, which is mediated by inactivation of PI3-K/Akt signaling. © 2015 Wiley Periodicals, Inc.

  3. PTEN/PI3K/Akt/VEGF signaling and the cross talk to KRIT1, CCM2, and PDCD10 proteins in cerebral cavernous malformations.

    PubMed

    Kar, Souvik; Samii, Amir; Bertalanffy, Helmut

    2015-04-01

    Cerebral cavernous malformations (CCM) are common vascular malformation of the brain and are associated with abnormal angiogenesis. Although the exact etiology and the underlying molecular mechanism are still under investigation, recent advances in the identification of the mutations in three genes and their interactions with different signaling pathways have shed light on our understanding of CCM pathogenesis. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is known to play a major role in angiogenesis. Studies have shown that the phosphatase and tensin homologue deleted on chromosome ten (PTEN), a tumor suppressor, is an antagonist regulator of the PI3K/Akt pathway and mediates angiogenesis by activating vascular endothelial growth factor (VEGF) expression. Here, we provide an update literature review on the current knowledge of the PTEN/PI3K/Akt/VEGF signaling in angiogenesis, more importantly in CCM pathogenesis. In addition to reviewing the current literatures, this article will also focus on the structural domain of the three CCM proteins and their interacting partners. Understanding the biology of these proteins with respect to their signaling counterpart will help to guide future research towards new therapeutic targets applicable for CCM treatment.

  4. Cyanidin-3-rutinoside increases glucose uptake by activating the PI3K/Akt pathway in 3T3-L1 adipocytes.

    PubMed

    Choi, Kyung Ha; Lee, Hyun Ah; Park, Mi Hwa; Han, Ji-Sook

    2017-09-01

    In this study, the effect of cyanidin-3-rutinoside (C3R) on glucose uptake by 3T3-L1 adipocytes was studied. C3R significantly increased glucose uptake, which was associated with enhanced plasma membrane glucose transporter type 4 (PM-GLUT4) expression in 3T3-L1 adipocytes. The potentiating effect of C3R on glucose uptake and PM-GLUT4 expression was related to enhanced phosphorylation of insulin receptor substrate 1 (IRS-1) and Akt, as well as augmented activation of phosphatidylinositol-3-kinase (PI3K) in the insulin signaling pathway. C3R induced glucose uptake was inhibited only by the PI3K inhibitor, but not by an AMPK inhibitor in 3T3-L1 adipocytes. Therefore, C3R likely up-regulates glucose uptake and PM-GLUT4 expression in 3T3-L1 adipocytes by activating the PI3K/Akt pathways. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. PI3K-GLUT4 Signal Pathway Associated with Effects of EX-B3 Electroacupuncture on Hyperglycemia and Insulin Resistance of T2DM Rats

    PubMed Central

    2016-01-01

    Objectives. To explore electroacupuncture's (EA's) effects on fasting blood glucose (FBG) and insulin resistance of type 2 diabetic mellitus (T2DM) model rats and give a possible explanation for the effects. Method. It takes high fat diet and intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) for model preparation. Model rats were randomly divided into T2DM Model group, EA weiwanxiashu (EX-B3) group, and sham EA group (n = 12/group). EA (2 Hz continuous wave, 2 mA, 20 min/day, 6 days/week, 4 weeks) was applied as intervention. FBG, area under curve (AUC) of oral glucose tolerance test (OGTT), insulin resistance index (HOMA-IR), pancreatic B cell function index (HOMA-B), skeletal muscle phosphorylated phosphatidylinositol-3-kinase (PI3K), glucose transporter 4 (GLUT4), and membrane GLUT4 protein expression were measured. Results. EA weiwanxiashu (EX-B3) can greatly upregulate model rat's significantly reduced skeletal muscle PI3K (Y607) and membrane GLUT4 protein expression (P < 0.01), effectively reducing model rats' FBG and AUC of OGTT (P < 0.01). The effects are far superior to sham EA group. Conclusion. EA weiwanxiashu (EX-B3) can upregulate skeletal muscle phosphorylated PI3K protein expression, to stimulate membrane translocation of GLUT4 and thereby increase skeletal muscle glucose intake to treat T2DM. PMID:27656242

  6. Modulation of phosphatidylinositol 3-kinase signaling reduces intimal hyperplasia in aortocoronary saphenous vein grafts.

    PubMed

    Hata, Jonathan A; Petrofski, Jason A; Schroder, Jacob N; Williams, Matthew L; Timberlake, Sarah H; Pippen, Anne; Corwin, Michael T; Solan, Amy K; Jakoi, Andre; Gehrig, Thomas R; Kontos, Christopher D; Milano, Carmelo A

    2005-06-01

    Fifty percent of human aortocoronary saphenous vein grafts are occluded after 10 years. Intimal hyperplasia is an initial step in graft occlusion and consists of vascular smooth muscle cell proliferation. Phosphatidylinositol 3-kinase and its downstream regulator, the inositol 3-phosphatase PTEN (phosphatase and tensin homolog deleted on chromosome 10), are important regulators of vascular smooth muscle cell proliferation, migration, and cell death. This study tests whether overexpression of PTEN in aortocoronary saphenous vein grafts can reduce intimal hyperplasia. Adult dogs underwent aortocoronary bypass grafting to the left anterior descending artery by using the autologous saphenous vein. Saphenous vein grafts were treated with phosphate-buffered saline (n = 9), empty adenovirus (n = 8), or adenovirus encoding for PTEN (n = 8). Arteriography at 30 and 90 days assessed saphenous vein graft patency. A subset received saphenous vein grafts treated with a marker transgene (beta-galactosidase, n = 3), empty adenovirus (n = 4), or adenovirus encoding for PTEN (n = 4) and were killed on postoperative day 3 to confirm expression. Vascular smooth muscle cells were isolated from canine saphenous vein infected with adenovirus encoding for PTEN, and immunoblotting and proliferation assays were performed. Saphenous vein graft transgene expression was confirmed by means of immunohistochemistry, immunoblotting, and polymerase chain reaction. Arteriograms revealed all saphenous vein grafts to be patent. Saphenous vein grafts treated with adenovirus encoding for PTEN demonstrated reduced intimal area compared with those treated with empty adenovirus and phosphate-buffered saline (1.39 +/- 0.11 vs 2.35 +/- 0.3 and 2.57 +/- 0.4 mm 2 , P < .05), and the intima/media ratio was lower in saphenous vein grafts treated with adenovirus encoding for PTEN (0.50 +/- 0.05 vs 1.43 +/- 0.18 and 1.11 +/- 0.14, P < .005). PTEN overexpression in vascular smooth muscle cells inhibited platelet

  7. PI3K/AKT/mTOR: role in breast cancer progression, drug resistance, and treatment.

    PubMed

    Guerrero-Zotano, Angel; Mayer, Ingrid A; Arteaga, Carlos L

    2016-12-01

    Anti-cancer cancer-targeted therapies are designed to exploit a particular vulnerability in the tumor, which in most cases results from its dependence on an oncogene and/or loss of a tumor suppressor. Mutations in the phosphoinositide 3-kinase (PI3K)/AKT/mTOR pathway are freqcuently found in breast cancers and associated with cellular transformation, tumorigenesis, cancer progression, and drug resistance. Several drugs targeting PI3K/ATK/mTOR are currently in clinical trials, mainly in combination with endocrine therapy and anti-HER2 therapy. These drugs are the focus of this review.

  8. The PI3K/AKT Pathway as a Target for Cancer Treatment.

    PubMed

    Mayer, Ingrid A; Arteaga, Carlos L

    2016-01-01

    Anticancer targeted therapies are designed to exploit a particular vulnerability in the tumor, which in most cases results from its dependence on an oncogene and/or loss of a tumor suppressor. Genes in the phosphoinositide 3-kinase (PI3K)/AKT pathway are the most frequently altered in human cancers. Aberrant activation of this pathway, as a result of these somatic alterations, is associated with cellular transformation, tumorigenesis, cancer progression, and drug resistance. Several drugs targeting PI3K/ATK are currently in clinical trials, alone or in combination, in both solid tumors and hematologic malignancies. These drugs are the focus of this review.

  9. Ovarian expressed microsomal epoxide hydrolase: Role in detoxification of 4-vinylcyclohexene diepoxide and regulation by phosphatidylinositol-3 kinase signaling

    SciTech Connect

    Bhattacharya, Poulomi; Sen, Nivedita; Hoyer, Patricia B.; Keating, Aileen F.

    2012-01-01

    4-vinylcyclohexene diepoxide (VCD) is a metabolite of 4-vinylcyclohexene (VCH) which has the potential to be formed in the ovary through CYP2E1 activity. VCD specifically destroys primordial and small primary follicles in the rodent ovary. Mouse ovaries exposed to VCD demonstrate increased mRNA and protein expression of microsomal epoxide hydrolase (mEH), and an inactive tetrol metabolite (4-(1,2-dihydroxy)ethyl-1,2-dihydroxycyclohexane) can be formed in mouse ovarian follicles, potentially through detoxification action of mEH. In contrast, mEH can bioactivate another ovotoxic chemical, 7,12-dimethylbenz[a]anthracene (DMBA) to a more toxic compound, DMBA-3,4-diol-1,2-epoxide. Thus, the present study evaluated a functional role for mEH during detoxification of VCD. Additionally, because inhibition of the phosphatidyinositol-3 kinase (PI3K) signaling pathway in a previous study protected primordial follicles from VCD-induced destruction, but accelerated DMBA-induced ovotoxicity, a role for PI3K in ovarian mEH regulation was evaluated. Using a post-natal day (PND) 4 Fischer 344 rat whole ovary culture system inhibition of mEH using cyclohexene oxide during VCD exposure resulted in a greater (P < 0.05) loss of primordial and small primary follicles relative to VCD-treated ovaries. Also, relative to controls, meh mRNA was increased (P < 0.05) on day 4 of VCD (30 μM) exposure, followed by increased (P < 0.05) mEH protein after 6 days. Furthermore, inhibition of PI3K signaling increased mEH mRNA and protein expression. Thus, these results support a functional role for mEH in the rat ovary, and demonstrate the involvement of PI3K signaling in regulation of ovarian xenobiotic metabolism by mEH. -- Highlights: ► Ovarian mEH functions to metabolize VCD to a less toxic compound. ► mEH expression is increased in a temporal pattern in response to VCD exposure. ► PI3K signaling is involved in regulation of ovarian mEH expression.

  10. Characterization of VPS34-IN1, a selective inhibitor of Vps34, reveals that the phosphatidylinositol 3-phosphate-binding SGK3 protein kinase is a downstream target of class III phosphoinositide 3-kinase.

    PubMed

    Bago, Ruzica; Malik, Nazma; Munson, Michael J; Prescott, Alan R; Davies, Paul; Sommer, Eeva; Shpiro, Natalia; Ward, Richard; Cross, Darren; Ganley, Ian G; Alessi, Dario R

    2014-11-01

    The Vps34 (vacuolar protein sorting 34) class III PI3K (phosphoinositide 3-kinase) phosphorylates PtdIns (phosphatidylinositol) at endosomal membranes to generate PtdIns(3)P that regulates membrane trafficking processes via its ability to recruit a subset of proteins possessing PtdIns(3)P-binding PX (phox homology) and FYVE domains. In the present study, we describe a highly selective and potent inhibitor of Vps34, termed VPS34-IN1, that inhibits Vps34 with 25 nM IC50 in vitro, but does not significantly inhibit the activity of 340 protein kinases or 25 lipid kinases tested that include all isoforms of class I as well as class II PI3Ks. Administration of VPS34-IN1 to cells induces a rapid dose-dependent dispersal of a specific PtdIns(3)P-binding probe from endosome membranes, within 1 min, without affecting the ability of class I PI3K to regulate Akt. Moreover, we