Long-term mTOR inhibitors administration evokes altered calcium homeostasis and platelet dysfunction in kidney transplant patients.

Long-term mTOR inhibitors administration evokes altered calcium homeostasis and platelet dysfunction in kidney transplant patients.

J Cell Mol Med. 2013 Apr 12;

Authors: L�pez E, Berna-Erro A, Bermejo N, Brull JM, Martinez R, Garcia Pino G, Alvarado R, Salido GM, Rosado JA, Cubero JJ, Redondo PC

Abstract
The use of the mammal target of rapamycin (mTOR) inhibitors has been consolidated as the therapy of election for preventing graft rejection in kidney transplant patients, despite their immunosuppressive activity is less strong than anti-calcineurin agents like tacrolimus and cyclosporine A. Furthermore, as mTOR is widely expressed, rapamycin (a macrolide antibiotic produced by Streptomyces hygroscopicus) is recommended in patients presenting neoplasia due to its antiproliferative actions. Hence, we have investigated whether rapamycin presents side effects in the physiology of other cell types different from leucocytes, such as platelets. Blood samples were drawn from healthy volunteers and kidney transplant patients long-term medicated with rapamycin: sirolimus and everolimus. Platelets were either loaded with fura-2 or directly stimulated, and immunoassayed or fixed with Laemmli's buffer to perform the subsequent analysis of platelet physiology. Our results indicate that rapamycin evokes a biphasic time-dependent alteration in calcium homeostasis and function in platelets from kidney transplant patients under rapamycin regime, as demonstrated by the reduction in granule secretion observed and subsequent impairment of platelet aggregation in these patients compared with healthy volunteers. Platelet count was also reduced in these patients, thus 41% of patients presented thrombocytopenia. All together our results show that long-term administration of rapamycin to kidney transplant patients evokes alteration in platelet function.

PMID: 23577651 [PubMed - as supplied by publisher]

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Multicenter Phase II Study of Tivozanib (AV-951) and Everolimus (RAD001) for Patients With Refractory, Metastatic Colorectal Cancer.

Multicenter Phase II Study of Tivozanib (AV-951) and Everolimus (RAD001) for Patients With Refractory, Metastatic Colorectal Cancer.

Oncologist. 2013 Apr 11;

Authors: Wolpin BM, Ng K, Zhu AX, Abrams T, Enzinger PC, McCleary NJ, Schrag D, Kwak EL, Allen JN, Bhargava P, Chan JA, Goessling W, Blaszkowsky LS, Supko JG, Elliot M, Sato K, Regan E, Meyerhardt JA, Fuchs CS

Abstract
BACKGROUND: Treatments that target the vascular endothelial growth factor (VEGF) pathway have efficacy in colorectal cancer. We evaluated tolerability and efficacy of tivozanib (an oral VEGF receptor-1, -2, -3 inhibitor) plus everolimus (an oral mammalian target of rapamycin inhibitor). METHODS: The phase Ib study followed a 3 + 3 dose-escalation design with three dose levels. The primary objective in the follow-on phase II study was improvement in 2-month progression-free survival (PFS) from 30% (historical benchmark) to 50% in patients with refractory, metastatic colorectal cancer. RESULTS: Dose-limiting toxicities in the phase Ib study were grade 3 fatigue and dehydration. Oral tivozanib (1 mg daily for 3 of 4 weeks) and oral everolimus (10 mg daily continuously) were advanced to a 40-patient phase II study. The most common grade 3-4 adverse events were thrombocytopenia and hypophosphatemia. The 2-month PFS rate was 50%, with 20 of 40 patients having stable disease (SD). Seven (18%) patients were treated for ?6 months. Median PFS and overall survival (OS) times were 3.0 months (95% confidence interval [CI]: 1.9-3.6 months) and 5.6 months (95% CI: 4.4-10.6 months), respectively. Patients who developed grade 1+ hypertension had increased SD rates (65.2% vs. 29.4%) and longer OS times (10.6 vs. 3.7 months). CONCLUSIONS: The oral combination of tivozanib and everolimus was well tolerated, with stable disease achieved in 50% of patients with refractory, metastatic colorectal cancer.

PMID: 23580238 [PubMed - as supplied by publisher]

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COX-2 in cancer: Gordian knot or Achilles heel?

COX-2 in cancer: Gordian knot or Achilles heel?

Front Pharmacol. 2013;4:34

Authors: Stasinopoulos I, Shah T, Penet MF, Krishnamachary B, Bhujwalla ZM

Abstract
The networks of blood and lymphatic vessels and of the extracellular matrix and their cellular and structural components, that are collectively termed the tumor microenvironment, are frequently co-opted and shaped by cancer cells to survive, invade, and form distant metastasis. With an enviable capacity to adapt to continually changing environments, cancer represents the epitome of functional chaos, a stark contrast to the hierarchical and organized differentiation processes that dictate the development and life of biological organisms. The consequences of changing landscapes such as hypoxia and acidic extracellular pH in and around tumors create a cascade of changes in multiple pathways and networks that become apparent only several years later as recurrence and metastasis. These molecular and phenotypic changes, several of which are mediated by COX-2, approach the complexities of a "Gordian Knot." We review evidence from our studies and from literature suggesting that cyclooxygenase-2 (COX-2) biology presents a nodal point in cancer biology and an "Achilles heel" of COX-2-dependent tumors.

PMID: 23579438 [PubMed - in process]

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Pharmacological Inhibition of Platelet-tumor Cell Cross-talk Prevents Platelet-induced Overexpression of Cyclooxygenase-2 in HT29 Human Colon Carcinoma Cells.

Pharmacological Inhibition of Platelet-tumor Cell Cross-talk Prevents Platelet-induced Overexpression of Cyclooxygenase-2 in HT29 Human Colon Carcinoma Cells.

Mol Pharmacol. 2013 Apr 11;

Authors: Dovizio M, Maier TJ, Alberti S, Di Francesco L, Marcantoni E, Munch G, John CM, Suess B, Sgambato A, Steinhilber D, Patrignani P

Abstract
Cyclooxygenase(COX)-2-derived prostanoids can influence several processes that are linked to carcinogenesis. We aimed to address the hypothesis that platelets contribute to aberrant COX-2 expression in HT29 colon carcinoma cells and to reveal the role of platelet-induced COX-2 on the expression of proteins involved in malignancy and marker genes of epithelial-mesenchymal transition(EMT). Human platelets co-cultured with HT29 cells rapidly adhered to cancer cells and induced COX-2 mRNA expression, but not protein synthesis which required the late release of platelet PDGF and COX-2 mRNA stabilization. Platelet-induced COX-2-dependent PGE2 synthesis in HT29 cells was involved in downregulation of p21(WAF1/CIP1) and upregulation of cyclinB1, since these effects were prevented by rofecoxib(a selective COX-2 inhibitor) and rescued by exogenous PGE2. Galectin-3, highly expressed in HT29 cells, is unique among galectins because it contains a collagen-like domain. Thus, we studied the role of galectin-3 and platelet collagen receptors in platelet-induced COX-2 overexpression. Inhibitors of galectin-3 function(?-lactose, a dominant-negative form of galectin-3,Gal-3C, and anti-galectin-3 antibody M3/38) or collagen receptor-mediated platelet adhesion(revacept, a dimeric collagen receptor GPVI-Fc) prevented aberrant COX-2 expression. Inhibition of platelet-cancer cell interaction by revacept was more effective than rofecoxib in preventing platelet-induced mRNA changes of EMT markers suggesting that direct cell-cell contact and aberrant COX-2 expression synergistically induced gene expression modifications associated with EMT. In conclusion, our findings provide the rationale for testing blockers of collagen binding sites, such as revacept, and galectin-3 inhibitors in the prevention of colon cancer metastasis in animal models followed by studies in patients.

PMID: 23580446 [PubMed - as supplied by publisher]

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[Celecoxib increased cellular ATRA sensitivity of human colon cancer cell lines through COX-2-independent mechanisms].

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[Celecoxib increased cellular ATRA sensitivity of human colon cancer cell lines through COX-2-independent mechanisms].

Yao Xue Xue Bao. 2009 Dec;44(12):1353-8

Authors: Liu JP, Wei HB, Zheng ZH, Guo WP, Fang JF

Abstract
Retinoid resistance has limited clinical activity of retinoids as differentiation-inducing and apoptosis-inducing drugs. The present study was designed to investigate whether celecoxib (selective COX-2 inhibitor) has effects on cellular retinoid sensitivity of human colon cancer cell lines and its possible mechanism. Cell viability was measured by MTT assay. Apoptosis was detected by Annexin-V/PI staining and flow cytometry assay. PGE2 production was measured by ELISA assay. Expression of RARbeta was assayed by Western blotting. The results showed that celecoxib enhanced the inhibitory effect of ATRA in both COX-2 high-expressing HT-29 and COX-2 low-expressing SW480 cell lines. Further study showed the ATRA and celecoxib combination induced greater apoptosis, and the addition of PGE2 did not affect the number of apoptotic cells induced by the combination. Moreover, NS398 (another selective COX-2 inhibitor) did not affect the inhibitory effects of ATRA on both cell lines. Western blotting showed that the expression of RARbeta in HT-29 cell lines increased in celecoxib group and combination group. And the addition of PGE2 did not affect the expression of RARbeta induced by celecoxib either. In conclusion, celecoxib increased expression of RARbeta and cellular ATRA sensitivity through COX-2-independent mechanisms, which may provide a potential strategy for combination therapy.

PMID: 21351468 [PubMed - indexed for MEDLINE]

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Angiotensin II Activates NF-?B Through AT1A Receptor Recruitment of ?-arrestin in Cultured Rat Vascular Smooth Muscle Cells.

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Angiotensin II Activates NF-?B Through AT1A Receptor Recruitment of ?-arrestin in Cultured Rat Vascular Smooth Muscle Cells.

Am J Physiol Cell Physiol. 2013 Apr 10;

Authors: Morinelli TA, Lee MH, Kendall RT, Luttrell L, Walker LP, Ullian ME

Abstract
Activation of the angiotensin II (AngII) AT1A receptor (AT1AR) in rat aorta vascular smooth muscle cells (RASMC) results in the increased synthesis of the pro-inflammatory enzyme cyclooxygenase 2 (COX-2). We have previously shown that nuclear localization of internalized AT1AR results in activation of transcription of the gene for COX-2, PTGS-2. Others have suggested that AngII stimulation of COX-2 protein synthesis is mediated by NF-?B. The purpose of the present study was to examine the interrelationship between AT1AR activation, ?-arrestin recruitment and NF-?B activation in the ability of AngII to increase COX-2 protein synthesis in RASMC. In the present study we utilized RASMC, inhibitors of the NF-?B pathway, ?-arrestin knock down, radioligand binding, immunoblotting and immunofluorescence to characterize the roles of AT1AR internalization, NF-?B activation and ?-arrestin in AngII-induced COX-2 synthesis. Ro 106-9920 or parthenolide, agents that inhibit the initial steps of NF-?B activation, blocked AngII-induced p65 NF-?B nuclear localization, COX-2 protein expression, ?-arrestin recruitment, and AT1AR internalization, without inhibiting AngII-induced p42/44 ERK activation. Curcumin, an inhibitor of NF-?B induced transcription, blocked AngII-induced COX-2 protein expression without altering AT1AR internalization, AngII-induced p65 NF-?B nuclear localization or p42/44 ERK activation. SiRNA-induced knockdown of ?-arrestin 1 and 2 inhibited AngII-induced p65 NF-?B nuclear localization. In vascular smooth muscle cells, internalization of the activated AT1AR mediated by ?-arrestins activates the NF-?B pathway, producing nuclear localization of the transcription factor and initiation of COX-2 protein synthesis; thereby linking the internalization of the receptor with the NF-?B pathway.

PMID: 23576578 [PubMed - as supplied by publisher]

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A preliminary study on the radiation-resistance mechanism in ovarian cancer.

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A preliminary study on the radiation-resistance mechanism in ovarian cancer.

J Cancer Res Ther. 2013 Jan-Mar;9(1):22-4

Authors: Liao Q, Zhang HM, Li HH, Zhou R, Mao HL, Chen YB, Cui MH

Abstract
Aim: The present study was designed to explore the radiation-resistance mechanism by interfering in checkpoints kinase 1 (CHK1) and DNA-activated protein kinase (DNA-PK) genes with short hairpin RNA (shRNA) transfection into Skov3 cells derived from ovarian cancer and HeLa cells derived from cervical cancer. Materials and Methods: The cultured Skov3 and HeLa cells were transfected with plasmid vectors containing CHK1 shRNA and DNA-PK shRNA, respectively, through Lipofectimine? 2000 mediation, and cultured for 20 hours before exposure to 2 Gy X-radiation. The cells were harvested 4 and 28 after X-irradiation respectively then washed 3 times with PBS. These cells were stained with Annexin V/PI and applied by flow cytometer to analyze alteration of apoptosis with software CellQuest. Results: The apoptotic response in Skov3 cells to X-radiation was significantly lower than that in HeLa cells at 4 hour (t = 15.22, P < 0.001) and 28 hours (t = 15.78, P < 0.001) of post-irradiation. The shRNA might not affect the apoptosis of Skov3 and HeLa cells, while shRNA-transfection significantly enhanced the apoptotic response in Skov3 cells to X-radiation as compared with that in HeLa cells. Conclusions: The present work suggests that the CHK1 and DNA-PK genes are very likely to play a role in developing a radiation resistance in ovarian cancer.

PMID: 23575069 [PubMed - in process]

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Cancer metabolism, stemness and tumor recurrence: MCT1 and MCT4 are functional biomarkers of metabolic symbiosis in head and neck cancer.

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Cancer metabolism, stemness and tumor recurrence: MCT1 and MCT4 are functional biomarkers of metabolic symbiosis in head and neck cancer.

Cell Cycle. 2013 Apr 10;12(9)

Authors: Curry JM, Tuluc M, Whitaker-Menezes D, Ames JA, Anantharam A, Oldt A, Leiby B, Cognetti DM, Sotgia F, Lisanti MP, Martinez-Outschoorn UE

Abstract
Here, we interrogated head and neck cancer (HNSCC) specimens (n = 12) to examine if different metabolic compartments (oxidative vs. glycolytic) co-exist in human tumors. A large panel of well-established biomarkers was employed to determine the metabolic state of proliferative cancer cells. Interestingly, cell proliferation in cancer cells, as marked by Ki-67 immunostaining, was strictly correlated with oxidative mitochondrial metabolism (OXPHOS) and the uptake of mitochondrial fuels, as detected via MCT1 expression (p < 0.001). More specifically, three metabolic tumor compartments were delineated: (1) proliferative and mitochondrial-rich cancer cells (Ki-67+/TOMM20+/COX+/MCT1+); (2) non-proliferative and mitochondrial-poor cancer cells (Ki-67-/TOMM20-/COX-/MCT1-); and (3) non-proliferative and mitochondrial-poor stromal cells (Ki-67-/TOMM20-/COX-/MCT1-). In addition, high oxidative stress (MCT4+) was very specific for cancer tissues. Thus, we next evaluated the prognostic value of MCT4 in a second independent patient cohort (n = 40). Most importantly, oxidative stress (MCT4+) in non-proliferating epithelial cancer cells predicted poor clinical outcome (tumor recurrence; p < 0.0001; log-rank test), and was functionally associated with FDG-PET avidity (p < 0.04). Similarly, oxidative stress (MCT4+) in tumor stromal cells was specifically associated with higher tumor stage (p < 0.03), and was a highly specific marker for cancer-associated fibroblasts (p < 0.001). We propose that oxidative stress is a key hallmark of tumor tissues that drives high-energy metabolism in adjacent proliferating mitochondrial-rich cancer cells, via the paracrine transfer of mitochondrial fuels (such as L-lactate and ketone bodies). New antioxidants and MCT4 inhibitors should be developed to metabolically target "three-compartment tumor metabolism" in head and neck cancers. It is remarkable that two "non-proliferating" populations of cells (Ki-67-/MCT4+) within the tumor can actually determine clinical outcome, likely by providing high-energy mitochondrial "fuels" for proliferative cancer cells to burn. Finally, we also show that in normal mucosal tissue, the basal epithelial "stem cell" layer is hyper-proliferative (Ki-67+), mitochondrial-rich (TOMM20+/COX+) and is metabolically programmed to use mitochondrial fuels (MCT1+), such as ketone bodies and L-lactate. Thus, oxidative mitochondrial metabolism (OXPHOS) is a common feature of both (1) normal stem cells and (2) proliferating cancer cells. As such, we should consider metabolically treating cancer patients with mitochondrial inhibitors (such as Metformin), and/or with a combination of MCT1 and MCT4 inhibitors, to target "metabolic symbiosis."

PMID: 23574725 [PubMed - as supplied by publisher]

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Optimizing megakaryocyte polyploidization by targeting multiple pathways of cytokinesis.

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Optimizing megakaryocyte polyploidization by targeting multiple pathways of cytokinesis.

Transfusion. 2012 Nov;52(11):2406-13

Authors: Avanzi MP, Chen A, He W, Mitchell WB

Abstract
BACKGROUND: Large-scale in vitro production of platelets (PLTs) from cord blood stem cells is one goal of stem cell research. One step toward this goal will be to produce polyploid megakaryocytes capable of releasing high numbers of PLTs. Megakaryocyte polyploidization requires distinct cytoskeletal and cellular mechanisms, including actin polymerization, myosin activation, microtubule formation, and increased DNA production. In this study we variably combined inhibition of these principal mechanisms of cytokinesis with the goal of driving polyploidization in megakaryocytes.
STUDY DESIGN AND METHODS: Megakaryocytes were derived from umbilical cord blood and cultured with reagents that inhibit distinct mechanisms of cytokinesis: Rho-Rock inhibitor (RRI), Src inhibitor (SI), nicotinamide (NIC), aurora B inhibitor (ABI), and myosin light chain kinase inhibitor (MLCKI). Combinations of reagents were used to determine their interactions and to maximize megakaryocyte ploidy.
RESULTS: Treatment with RRI, NIC, SI, and ABI, but not with MLCKI, increased the final ploidy and RRI was the most effective single reagent. RRI and MLCKI, both inhibitors of MLC activation, resulted in opposite ploidy outcomes. Combinations of reagents also increased ploidy and the use of NIC, SI, and ABI was as effective as RRI alone. Addition of MLCKI to NIC, SI, and ABI reached the highest level of polyploidization.
CONCLUSION: Megakaryocyte polyploidization results from modulation of a combination of distinct cytokinesis pathways. Reagents targeting distinct cytoskeletal pathways produced additive effects in final megakaryocyte ploidy. The RRI, however, showed no additive effect but produced a high final ploidy due to overlapping inhibition of multiple cytokinesis pathways.

PMID: 22612069 [PubMed - indexed for MEDLINE]

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Toll-like receptor 9 agonist IMO cooperates with everolimus in renal cell carcinoma by interfering with tumour growth and angiogenesis.

Toll-like receptor 9 agonist IMO cooperates with everolimus in renal cell carcinoma by interfering with tumour growth and angiogenesis.

Br J Cancer. 2013 Apr 9;

Authors: Damiano V, Rosa R, Formisano L, Nappi L, Gelardi T, Marciano R, Cozzolino I, Troncone G, Agrawal S, Veneziani BM, De Placido S, Bianco R, Tortora G

Abstract
Background:Targeting the mammalian target of rapamycin by everolimus is a successful approach for renal cell carcinoma (RCC) therapy. The Toll-like receptor 9 agonist immune modulatory oligonucleotide (IMO) exhibits direct antitumour and antiangiogenic activity and cooperates with both epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF) inhibitors.Methods:We tested the combination of IMO and everolimus on models of human RCC with different Von-Hippel Lindau (VHL) gene status, both in vitro and in nude mice. We studied their direct antiangiogenic effects on human umbilical vein endothelial cells.Results:Both IMO and everolimus inhibited in vitro growth and survival of RCC cell lines, and their combination produced a synergistic inhibitory effect. Moreover, everolimus plus IMO interfered with EGFR-dependent signaling and reduced VEGF secretion in both VHL wild-type and mutant cells. In RCC tumour xenografts, IMO plus everolimus caused a potent and long-lasting cooperative antitumour activity, with reduction of tumour growth, prolongation of mice survival and inhibition of signal transduction. Furthermore, IMO and everolimus impaired the main endothelial cell functions.Conclusion:A combined treatment with everolimus and IMO is effective in VHL wild-type and mutant models of RCC by interfering with tumour growth and angiogenesis, thus representing a potentially effective, rationale-based combination to be translated in the clinical setting.British Journal of Cancer advance online publication, 9 April 2013; doi:10.1038/bjc.2013.153 www.bjcancer.com.

PMID: 23571736 [PubMed - as supplied by publisher]

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Toll-like receptor 9 agonist IMO cooperates with everolimus in renal cell carcinoma by interfering with tumour growth and angiogenesis.

Toll-like receptor 9 agonist IMO cooperates with everolimus in renal cell carcinoma by interfering with tumour growth and angiogenesis.

Br J Cancer. 2013 Apr 9;

Authors: Damiano V, Rosa R, Formisano L, Nappi L, Gelardi T, Marciano R, Cozzolino I, Troncone G, Agrawal S, Veneziani BM, De Placido S, Bianco R, Tortora G

Abstract
Background:Targeting the mammalian target of rapamycin by everolimus is a successful approach for renal cell carcinoma (RCC) therapy. The Toll-like receptor 9 agonist immune modulatory oligonucleotide (IMO) exhibits direct antitumour and antiangiogenic activity and cooperates with both epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF) inhibitors.Methods:We tested the combination of IMO and everolimus on models of human RCC with different Von-Hippel Lindau (VHL) gene status, both in vitro and in nude mice. We studied their direct antiangiogenic effects on human umbilical vein endothelial cells.Results:Both IMO and everolimus inhibited in vitro growth and survival of RCC cell lines, and their combination produced a synergistic inhibitory effect. Moreover, everolimus plus IMO interfered with EGFR-dependent signaling and reduced VEGF secretion in both VHL wild-type and mutant cells. In RCC tumour xenografts, IMO plus everolimus caused a potent and long-lasting cooperative antitumour activity, with reduction of tumour growth, prolongation of mice survival and inhibition of signal transduction. Furthermore, IMO and everolimus impaired the main endothelial cell functions.Conclusion:A combined treatment with everolimus and IMO is effective in VHL wild-type and mutant models of RCC by interfering with tumour growth and angiogenesis, thus representing a potentially effective, rationale-based combination to be translated in the clinical setting.British Journal of Cancer advance online publication, 9 April 2013; doi:10.1038/bjc.2013.153 www.bjcancer.com.

PMID: 23571736 [PubMed - as supplied by publisher]

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Optimizing megakaryocyte polyploidization by targeting multiple pathways of cytokinesis.

Related Articles

Optimizing megakaryocyte polyploidization by targeting multiple pathways of cytokinesis.

Transfusion. 2012 Nov;52(11):2406-13

Authors: Avanzi MP, Chen A, He W, Mitchell WB

Abstract
BACKGROUND: Large-scale in vitro production of platelets (PLTs) from cord blood stem cells is one goal of stem cell research. One step toward this goal will be to produce polyploid megakaryocytes capable of releasing high numbers of PLTs. Megakaryocyte polyploidization requires distinct cytoskeletal and cellular mechanisms, including actin polymerization, myosin activation, microtubule formation, and increased DNA production. In this study we variably combined inhibition of these principal mechanisms of cytokinesis with the goal of driving polyploidization in megakaryocytes.
STUDY DESIGN AND METHODS: Megakaryocytes were derived from umbilical cord blood and cultured with reagents that inhibit distinct mechanisms of cytokinesis: Rho-Rock inhibitor (RRI), Src inhibitor (SI), nicotinamide (NIC), aurora B inhibitor (ABI), and myosin light chain kinase inhibitor (MLCKI). Combinations of reagents were used to determine their interactions and to maximize megakaryocyte ploidy.
RESULTS: Treatment with RRI, NIC, SI, and ABI, but not with MLCKI, increased the final ploidy and RRI was the most effective single reagent. RRI and MLCKI, both inhibitors of MLC activation, resulted in opposite ploidy outcomes. Combinations of reagents also increased ploidy and the use of NIC, SI, and ABI was as effective as RRI alone. Addition of MLCKI to NIC, SI, and ABI reached the highest level of polyploidization.
CONCLUSION: Megakaryocyte polyploidization results from modulation of a combination of distinct cytokinesis pathways. Reagents targeting distinct cytoskeletal pathways produced additive effects in final megakaryocyte ploidy. The RRI, however, showed no additive effect but produced a high final ploidy due to overlapping inhibition of multiple cytokinesis pathways.

PMID: 22612069 [PubMed - indexed for MEDLINE]

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Reduction in first and recurrent cardiovascular events with ticagrelor compared with clopidogrel in the PLATO Study.

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Reduction in first and recurrent cardiovascular events with ticagrelor compared with clopidogrel in the PLATO Study.

Circulation. 2013 Feb 12;127(6):673-80

Authors: Kohli P, Wallentin L, Reyes E, Horrow J, Husted S, Angiolillo DJ, Ardissino D, Maurer G, Morais J, Nicolau JC, Oto A, Storey RF, James SK, Cannon CP

Abstract
BACKGROUND: We sought to evaluate the effect of potent platelet inhibition after acute coronary syndrome on total (ie, first and recurrent) occurrences of any of the primary outcome events (e.g., cardiovascular death, myocardial infarction, and stroke) as well as on other ischemic events, such as urgent revascularization, (severe) recurrent ischemia, transient ischemic attacks, and arterial thrombotic events.
METHODS AND RESULTS: In the PLATelet inhibition and patient Outcomes (PLATO) study, 18 624 patients presenting with acute coronary syndromes randomly received ticagrelor (n=9333) or clopidogrel (n=9291). Cox proportional hazard models were used to calculate time to first event and hazard ratios. Total events were compared using a Poisson regression model, and time to second event or death was calculated with the Wei Lin Weissfeld method. Patients randomized to ticagrelor had 1057 total primary end point events versus 1225 for patients on clopidogrel (rate ratio, 0.86; 95% confidence interval, 0.79-0.93; P=0.003). The number of additional events was numerically lower for ticagrelor (189 versus 205; P=0.40), resulting in a hazard for time to second event/death of 0.80 (95% confidence interval, 0.70-0.90; P<0.001) and a number needed to treat of 54. For cardiovascular death/myocardial infarction/stroke/(severe) recurrent ischemia/transient ischemic attack/arterial thrombotic events, total events were fewer with ticagrelor (2030 versus 2290; rate ratio, 0.88; 95% confidence interval, 0.82-0.95; P<0.001), with fewer recurrent events with ticagrelor (740 versus 834; P=0.01) and a highly significant concurrent reduction in hazard for time to second event or death of 0.83 (95% confidence interval, 0.75-0.91; P<0.001). Recurrent PLATO major or Thrombolysis in Myocardial Infarction (TIMI) major non-coronary artery bypass graft bleeding events were infrequent and not different between the two therapies (P=0.96 and 0.38, respectively).
CONCLUSIONS: In PLATO, treatment with ticagrelor compared with clopidogrel resulted in a reduction in total events, including first and subsequent recurrent cardiovascular events, when compared with clopidogrel. These types of analyses demonstrate an even greater absolute benefit of ticagrelor over clopidogrel than previously reported.
CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov/. Unique identifier: NCT00391872.

PMID: 23277305 [PubMed - indexed for MEDLINE]

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Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-?/BMP signaling and orphan nuclear receptors.

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Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-?/BMP signaling and orphan nuclear receptors.

PLoS One. 2012;7(7):e40255

Authors: Boulanger A, Farge M, Ramanoudjame C, Wharton K, Dura JM

Abstract
Larval motor neurons remodel during Drosophila neuro-muscular junction dismantling at metamorphosis. In this study, we describe the motor neuron retraction as opposed to degeneration based on the early disappearance of ?-Spectrin and the continuing presence of Tubulin. By blocking cell dynamics with a dominant-negative form of Dynamin, we show that phagocytes have a key role in this process. Importantly, we show the presence of peripheral glial cells close to the neuro-muscular junction that retracts before the motor neuron. We show also that in muscle, expression of EcR-B1 encoding the steroid hormone receptor required for postsynaptic dismantling, is under the control of the ftz-f1/Hr39 orphan nuclear receptor pathway but not the TGF-? signaling pathway. In the motor neuron, activation of EcR-B1 expression by the two parallel pathways (TGF-? signaling and nuclear receptor) triggers axon retraction. We propose that a signal from a TGF-? family ligand is produced by the dismantling muscle (postsynapse compartment) and received by the motor neuron (presynaptic compartment) resulting in motor neuron retraction. The requirement of the two pathways in the motor neuron provides a molecular explanation for the instructive role of the postsynapse degradation on motor neuron retraction. This mechanism insures the temporality of the two processes and prevents motor neuron pruning before postsynaptic degradation.

PMID: 22792255 [PubMed - indexed for MEDLINE]

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Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Mol Cancer Ther. 2013 Feb 26;

Authors: Konecny GE, Kolarova T, O'Brien NA, Winterhoff B, Yang G, Qi J, Qi Z, Venkatesan N, Ayala R, Luo T, Finn RS, Kristof J, Galderisi C, Graus Porta D, Anderson L, Shi MM, Yovine A, Slamon DJ

Abstract
The recent identification of activating fibroblast growth factor receptor 2 (FGFR2) mutations in endometrial cancer (EC) has generated an opportunity for a novel target-based therapy. Here we explore the therapeutic potential of two FGFR inhibitors, the multi-kinase inhibitor dovitinib (TKI258) and the more selective FGFR inhibitor NVP-BGJ398 for the treatment of EC. We examined the effects of both inhibitors on tumor cell growth, FGFR2 signaling, cell cycle and apoptosis using a panel of 20 molecularly characterized human EC cell lines. Anchorage independent growth was studied using soft agar assays. In vivo studies were conducted using EC xenograft models. Cell lines with activating FGFR2 mutations (S252W, N550K) were more sensitive to dovitinib or NVP-BGJ398 when compared to their FGFR2 wild-type counterparts (p=0.073 and p=0.021, respectively). Both agents inhibited FGFR2 signaling, induced cell cycle arrest and significantly increased apoptosis in FGFR2 mutant lines. In vitro, dovitinib and NVP-BGJ398 were both potent at inhibiting cell growth of FGFR2 mutant EC cells but the activity of dovitinib was less restricted to FGFR2 mutant lines when compared to NVP-BGJ398. In vivo, dovitinib and NVP-BGJ398 significantly inhibited the growth of FGFR2 mutated EC xenograft models. In addition, dovitinib showed significant antitumor activity in FGFR2 wild-type EC xenograft models including complete tumor regressions in a long term in vivo study. Dovitinib and NVP-BGJ398 warrant further clinical evaluation in patients with FGFR2 mutated EC. Dovitinib may have antitumor activity in EC beyond FGFR2 mutated cases and may permit greater flexibility in patient selection.

PMID: 23443805 [PubMed - as supplied by publisher]

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Image-based high-throughput screening for inhibitors of angiogenesis.

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Image-based high-throughput screening for inhibitors of angiogenesis.

Methods Mol Biol. 2013;931:139-51

Authors: Evensen L, Link W, Lorens JB

Abstract
Automated multicolor fluorescence microscopy facilitates high-throughput quantitation of cellular parameters of complex, organotypic systems. In vitro co-cultured vascular cells form capillary-like networks that model facets of angiogenesis, making it an attractive alternative for anti-angiogenic drug discovery. We have adapted this angiogenesis assay system to a high-throughput format to enable automated image-based high-throughput screening of live primary human vascular cell co-cultures with chemical libraries for anti-angiogenic drug discovery. Protocols are described for setup of a fluorescence-based co-culture assay, live cell image acquisition, image analysis of morphological parameters, and screening data handling.

PMID: 23027002 [PubMed - indexed for MEDLINE]

CDK5 inhibitor chk2 inhibitor selleck chemical checkpoint inhibitors selleckchem

Use of aspirin, but not other non-steroidal anti-inflammatory drugs is associated with decreased prostate cancer risk at the population level.

Related Articles

Use of aspirin, but not other non-steroidal anti-inflammatory drugs is associated with decreased prostate cancer risk at the population level.

Eur J Cancer. 2013 Mar;49(4):938-45

Authors: Veitonm�ki T, Tammela TL, Auvinen A, Murtola TJ

Abstract
The cyclooxygenase 2 (COX-2) enzyme overexpression in prostate cancer has led to the hypothesis that COX-2 inhibition may reduce prostate cancer growth. Some previous studies have linked the usage of COX-2 inhibiting non-steroidal anti-inflammatory drugs (NSAIDs) with a decreased prostate cancer risk. We estimated the association between cumulative COX-2 inhibition by NSAID usage and prostate cancer risk at population level. All new prostate cancer cases in Finland during 1995-2002 and matched controls (24,657 case-control pairs) were identified from national registries. Detailed information on medication purchases was obtained from a national prescription database. A total cumulative COX-2 inhibition value was calculated based on total cumulative mg amount of each NSAID drug and the drug-specific COX-1/COX-2 inhibition ratio. Prostate cancer risk was analysed with propensity score-matched conditional logistic regression model. In total, 53.8% of the cases and 46.5% of the controls had any prescription-use of NSAIDs, while 8.1% and 7.9%, respectively, had used aspirin. Compared to the non-users, any NSAID use was associated with an elevated overall prostate cancer risk (46.4% versus 53.6%, respectively; odds ratio [OR] 1.3, 95% confidence interval [CI] 1.3, 1.4) and risk of advanced cancer (11.8% versus 14.1%; OR 1.6, 95% CI 1.5, 1.8). The risk remained elevated despite the amount of cumulative COX-2 inhibition. In a separate analysis, the risk increase was similar for each NSAID with the exception of aspirin, which was associated with a decreased overall prostate cancer risk (OR 0.90, 95% CI 0.84, 0.96) in a dose-dependent fashion. NSAID use is associated with an increased prostate cancer risk at the population level regardless of the COX-2 inhibition. This may be explained by systematic differences between prescription NSAID users and non-users. In contrast, aspirin use is associated with a decreased overall prostate cancer risk. Further studies on aspirin and prostate cancer will be needed.

PMID: 23079475 [PubMed - indexed for MEDLINE]

Caspase inhibitors selleck Caspase-9 inhibitor selleck chemicals reversible Caspase inhibitor selleck chemicals

Image-based high-throughput screening for inhibitors of angiogenesis.

Related Articles

Image-based high-throughput screening for inhibitors of angiogenesis.

Methods Mol Biol. 2013;931:139-51

Authors: Evensen L, Link W, Lorens JB

Abstract
Automated multicolor fluorescence microscopy facilitates high-throughput quantitation of cellular parameters of complex, organotypic systems. In vitro co-cultured vascular cells form capillary-like networks that model facets of angiogenesis, making it an attractive alternative for anti-angiogenic drug discovery. We have adapted this angiogenesis assay system to a high-throughput format to enable automated image-based high-throughput screening of live primary human vascular cell co-cultures with chemical libraries for anti-angiogenic drug discovery. Protocols are described for setup of a fluorescence-based co-culture assay, live cell image acquisition, image analysis of morphological parameters, and screening data handling.

PMID: 23027002 [PubMed - indexed for MEDLINE]

checkpoint inhibitors selleck chemical chk2 inhibitor selleckchem checkpoint inhibitors selleck chemicals

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Mol Cancer Ther. 2013 Feb 26;

Authors: Konecny GE, Kolarova T, O'Brien NA, Winterhoff B, Yang G, Qi J, Qi Z, Venkatesan N, Ayala R, Luo T, Finn RS, Kristof J, Galderisi C, Graus Porta D, Anderson L, Shi MM, Yovine A, Slamon DJ

Abstract
The recent identification of activating fibroblast growth factor receptor 2 (FGFR2) mutations in endometrial cancer (EC) has generated an opportunity for a novel target-based therapy. Here we explore the therapeutic potential of two FGFR inhibitors, the multi-kinase inhibitor dovitinib (TKI258) and the more selective FGFR inhibitor NVP-BGJ398 for the treatment of EC. We examined the effects of both inhibitors on tumor cell growth, FGFR2 signaling, cell cycle and apoptosis using a panel of 20 molecularly characterized human EC cell lines. Anchorage independent growth was studied using soft agar assays. In vivo studies were conducted using EC xenograft models. Cell lines with activating FGFR2 mutations (S252W, N550K) were more sensitive to dovitinib or NVP-BGJ398 when compared to their FGFR2 wild-type counterparts (p=0.073 and p=0.021, respectively). Both agents inhibited FGFR2 signaling, induced cell cycle arrest and significantly increased apoptosis in FGFR2 mutant lines. In vitro, dovitinib and NVP-BGJ398 were both potent at inhibiting cell growth of FGFR2 mutant EC cells but the activity of dovitinib was less restricted to FGFR2 mutant lines when compared to NVP-BGJ398. In vivo, dovitinib and NVP-BGJ398 significantly inhibited the growth of FGFR2 mutated EC xenograft models. In addition, dovitinib showed significant antitumor activity in FGFR2 wild-type EC xenograft models including complete tumor regressions in a long term in vivo study. Dovitinib and NVP-BGJ398 warrant further clinical evaluation in patients with FGFR2 mutated EC. Dovitinib may have antitumor activity in EC beyond FGFR2 mutated cases and may permit greater flexibility in patient selection.

PMID: 23443805 [PubMed - as supplied by publisher]

ATP-competitive Caspase inhibitor selleckchem CDK5 inhibitor chk2 inhibitor selleck chemical

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Mol Cancer Ther. 2013 Feb 26;

Authors: Konecny GE, Kolarova T, O'Brien NA, Winterhoff B, Yang G, Qi J, Qi Z, Venkatesan N, Ayala R, Luo T, Finn RS, Kristof J, Galderisi C, Graus Porta D, Anderson L, Shi MM, Yovine A, Slamon DJ

Abstract
The recent identification of activating fibroblast growth factor receptor 2 (FGFR2) mutations in endometrial cancer (EC) has generated an opportunity for a novel target-based therapy. Here we explore the therapeutic potential of two FGFR inhibitors, the multi-kinase inhibitor dovitinib (TKI258) and the more selective FGFR inhibitor NVP-BGJ398 for the treatment of EC. We examined the effects of both inhibitors on tumor cell growth, FGFR2 signaling, cell cycle and apoptosis using a panel of 20 molecularly characterized human EC cell lines. Anchorage independent growth was studied using soft agar assays. In vivo studies were conducted using EC xenograft models. Cell lines with activating FGFR2 mutations (S252W, N550K) were more sensitive to dovitinib or NVP-BGJ398 when compared to their FGFR2 wild-type counterparts (p=0.073 and p=0.021, respectively). Both agents inhibited FGFR2 signaling, induced cell cycle arrest and significantly increased apoptosis in FGFR2 mutant lines. In vitro, dovitinib and NVP-BGJ398 were both potent at inhibiting cell growth of FGFR2 mutant EC cells but the activity of dovitinib was less restricted to FGFR2 mutant lines when compared to NVP-BGJ398. In vivo, dovitinib and NVP-BGJ398 significantly inhibited the growth of FGFR2 mutated EC xenograft models. In addition, dovitinib showed significant antitumor activity in FGFR2 wild-type EC xenograft models including complete tumor regressions in a long term in vivo study. Dovitinib and NVP-BGJ398 warrant further clinical evaluation in patients with FGFR2 mutated EC. Dovitinib may have antitumor activity in EC beyond FGFR2 mutated cases and may permit greater flexibility in patient selection.

PMID: 23443805 [PubMed - as supplied by publisher]

CDK5 inhibitor chk2 inhibitor selleck chemical checkpoint inhibitors selleckchem

Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-?/BMP signaling and orphan nuclear receptors.

Related Articles

Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-?/BMP signaling and orphan nuclear receptors.

PLoS One. 2012;7(7):e40255

Authors: Boulanger A, Farge M, Ramanoudjame C, Wharton K, Dura JM

Abstract
Larval motor neurons remodel during Drosophila neuro-muscular junction dismantling at metamorphosis. In this study, we describe the motor neuron retraction as opposed to degeneration based on the early disappearance of ?-Spectrin and the continuing presence of Tubulin. By blocking cell dynamics with a dominant-negative form of Dynamin, we show that phagocytes have a key role in this process. Importantly, we show the presence of peripheral glial cells close to the neuro-muscular junction that retracts before the motor neuron. We show also that in muscle, expression of EcR-B1 encoding the steroid hormone receptor required for postsynaptic dismantling, is under the control of the ftz-f1/Hr39 orphan nuclear receptor pathway but not the TGF-? signaling pathway. In the motor neuron, activation of EcR-B1 expression by the two parallel pathways (TGF-? signaling and nuclear receptor) triggers axon retraction. We propose that a signal from a TGF-? family ligand is produced by the dismantling muscle (postsynapse compartment) and received by the motor neuron (presynaptic compartment) resulting in motor neuron retraction. The requirement of the two pathways in the motor neuron provides a molecular explanation for the instructive role of the postsynapse degradation on motor neuron retraction. This mechanism insures the temporality of the two processes and prevents motor neuron pruning before postsynaptic degradation.

PMID: 22792255 [PubMed - indexed for MEDLINE]

checkpoint inhibitors selleckchem pan Chk inhibitor selleck chemicals checkpoint inhibitors selleck chemical

EMD 1214063 and EMD 1204831 constitute a new class of potent and highly selective c-Met inhibitors.

EMD 1214063 and EMD 1204831 constitute a new class of potent and highly selective c-Met inhibitors.

Clin Cancer Res. 2013 Apr 3;

Authors: Bladt F, Faden B, Friese-Hamim M, Knuehl C, Wilm C, Frittschen C, Graedler U, Meyring M, Dorsch D, Jaehrling F, Pehl U, Stieber F, Schadt O, Blaukat A

Abstract
PURPOSE: The mesenchymal-epithelial transition factor (c-Met) receptor, also known as hepatocyte growth factor receptor (HGFR), controls morphogenesis, a process that is physiologically required for embryonic development and tissue repair. Aberrant c-Met activation is associated with a variety of human malignancies including cancers of the lung, kidney, stomach, liver, and brain. In this study, we investigated the properties of two novel compounds, developed to selectively inhibit the c-Met receptor in anti-tumor therapeutic interventions. EXPERIMENTAL DESIGN: The pharmacologic properties, c-Met inhibitory activity, and anti-tumor effects of EMD 1214063 and EMD 1204831 were investigated in vitro and in vivo, using human cancer cell lines and mouse xenograft models. RESULTS: EMD 1214063 and EMD 1204831 selectively suppressed the c-Met receptor tyrosine kinase activity. Their inhibitory activity was potent (inhibitory 50% concentration [IC50], 3 nM and 9 nM, respectively) and highly selective, when compared to their effect on a panel of 242 human kinases. Both EMD 1214063 and EMD 1204831 inhibited c-Met phosphorylation and downstream signaling in a dose-dependent fashion, but differed in the duration of their inhibitory activity. In murine xenograft models, both compounds induced regression of human tumors, regardless of whether c-Met activation was HGF dependent or independent. Both drugs were well tolerated and induced no substantial weight loss after >3 weeks of treatment. CONCLUSIONS: Our results indicate selective c-Met inhibition by EMD 1214063 and EMD 1204831 and strongly support clinical testing of these compounds in the context of molecularly targeted anti-cancer strategies.

PMID: 23553846 [PubMed - as supplied by publisher]

rad001 ecdysone chir-258

DNA-PK phosphorylation of IGFBP-3 is required to Prevent Apoptosis in Retinal Endothelial cells cultured in High Glucose.

DNA-PK phosphorylation of IGFBP-3 is required to Prevent Apoptosis in Retinal Endothelial cells cultured in High Glucose.

Invest Ophthalmol Vis Sci. 2013 Apr 4;

Authors: Zhang Q, Steinle JJ

Abstract
PURPOSE: The goal of this study was to determine whether Compound 49b stimulates insulin-like growth factor binding protein-3 (IGFBP-3) activation in retinal endothelial cells (REC) through DNA-dependent protein kinase (DNA-PK). METHODS: REC were grown in normal glucose (5 mM) or high glucose medium (25 mM). Some cells were transfected with protein kinase A (PKA) siRNA, following treatment with 50 nM Compound 49b, a novel ?-adrenergic receptor agonist. Cell proteins were extracted and analyzed for DNA-PK expression by Western blotting. Additional cells were treated with or without NU7441 (a specific DNA-PK inhibitor) prior to Compound 49b treatment. Cell lysates were processed for IGFBP-3 ELISA analyses and Western blotting to measure casein kinase 2 (CK2). Immunoprecipitation for total and phospho-IGFBP-3 and cell death measurements were done after transfection with the S156A IGFBP-3 mutation (key phosphorylation site involved in DNA-PK) plasmid DNA. RESULTS: Compound 49b required DNA-PK to activate IGFBP-3 in REC. IGFBP-3 activation was significantly reduced following treatment with either the DNA-PK inhibitor or following transfection with the IGFBP-3 S156A mutant plasmid (p < 0.05). Significant increases in cell death were also observed in cells transfected with the IGFBP-3 S156A mutant plasmid (p < 0.05). Casein kinase levels were not altered after treatment with NU7741 or Compound 49b. CONCLUSIONS: Our finding suggested Compound 49b induces DNA-PK levels through PKA activity. DNA-PK is required for Compound 49b-induced activation of IGFBP-3, leading to inhibition of REC cell death.

PMID: 23557743 [PubMed - as supplied by publisher]

ecdysone chir-258 dovitinib

Saucerneol D inhibits eicosanoid generation and degranulation through suppression of Syk kinase in mast cells.

Related Articles

Saucerneol D inhibits eicosanoid generation and degranulation through suppression of Syk kinase in mast cells.

Food Chem Toxicol. 2012 Dec;50(12):4382-8

Authors: Lu Y, Li Y, Seo CS, Murakami M, Son JK, Chang HW

Abstract
Previously we reported that saucerneol D (SD), a naturally occurring sesquilignan isolated from Saururus chinensis (S. chinensis) suppressed lipopolysaccharide (LPS)-induced nitric oxide (NO) production in RAW 264.7 cells. The aim of this study was to elucidate whether SD modulates the generation of other inflammatory mediators in activated mast cells. We investigated the effects of SD on cyclooxygenase-2 (COX-2)-dependent prostaglandin D(2) (PGD(2)) and 5-lipoxygenase (5-LO)-dependent leukotriene C(4) (LTC(4)) generations as well as degranulation in cytokine-stimulated mouse bone marrow-derived mast cells (BMMCs). Biochemical analyses of the cytokine-mediated signaling pathways showed that SD suppressed the phosphorylation of Syk kinase and multiple downstream signaling processes including phospholipase C?1 (PLC?1)-mediated intracellular Ca(2+) influx and activation of mitogen-activated protein kinases (MAPKs; including extracellular signal-regulated kinase 1/2 (ERK1/2), c-Jun NH(2)-terminal kinase (JNK) and p38) and the nuclear factor-?B (NF-?B) pathway. Taken together, the present study suggests that SD suppresses eicosanoid generation and degranulation through Syk-dependent pathway in BMMCs.

PMID: 22982805 [PubMed - indexed for MEDLINE]

chir-258 dovitinib dna-pk

The short form of RON is expressed in acute myeloid leukemia and sensitizes leukemic cells to cMET inhibitors.

Related Articles

The short form of RON is expressed in acute myeloid leukemia and sensitizes leukemic cells to cMET inhibitors.

Leukemia. 2013 Feb;27(2):325-35

Authors: Fialin C, Larrue C, Vergez F, Sarry JE, Bertoli S, Mansat-De Mas V, Demur C, Delabesse E, Payrastre B, Manenti S, Roche S, R�cher C

Abstract
Several receptor tyrosine kinases (TKs) are involved in the pathogenesis of acute myeloid leukemia (AML). Here, we have assessed the expression of the Recepteur d'Origine Nantais (RON) in leukemic cell lines and samples from AML patients. In a series of 86 AML patients, we show that both the full length and/or the short form (sf) of RON are expressed in 51% and 43% of cases, respectively. Interestingly, sfRON is not expressed in normal CD34+ hematopoietic cells and induces part of its oncogenic signaling through interaction with the Src kinase Lyn. sfRON-mediated signaling in leukemic cells also involves mTORC1, the proapoptotic bcl2-family member, BAD, but not the phosphatidylinositol 3-kinase/Akt pathway. Furthermore, the expression of sfRON was specifically downregulated by 5-azacytidine (AZA). Conversely, AZA could induce the expression of sfRON in sfRON-negative leukemic cells suggesting that the activity of this drug in AML and myelodysplastic syndromes could involve modulation of TKs. cMET/RON inhibitors exhibited an antileukemic activity exclusively in AML samples and cell lines expressing sfRON. These results might support clinical trials evaluating cMET/RON inhibitors in AML patients expressing sfRON.

PMID: 22902361 [PubMed - indexed for MEDLINE]

chir-258 dovitinib dna-pk

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Activity of the Fibroblast Growth Factor Receptor Inhibitors Dovitinib (TKI258) and NVP-BGJ398 in Human Endometrial Cancer Cells.

Mol Cancer Ther. 2013 Feb 26;

Authors: Konecny GE, Kolarova T, O'Brien NA, Winterhoff B, Yang G, Qi J, Qi Z, Venkatesan N, Ayala R, Luo T, Finn RS, Kristof J, Galderisi C, Graus Porta D, Anderson L, Shi MM, Yovine A, Slamon DJ

Abstract
The recent identification of activating fibroblast growth factor receptor 2 (FGFR2) mutations in endometrial cancer (EC) has generated an opportunity for a novel target-based therapy. Here we explore the therapeutic potential of two FGFR inhibitors, the multi-kinase inhibitor dovitinib (TKI258) and the more selective FGFR inhibitor NVP-BGJ398 for the treatment of EC. We examined the effects of both inhibitors on tumor cell growth, FGFR2 signaling, cell cycle and apoptosis using a panel of 20 molecularly characterized human EC cell lines. Anchorage independent growth was studied using soft agar assays. In vivo studies were conducted using EC xenograft models. Cell lines with activating FGFR2 mutations (S252W, N550K) were more sensitive to dovitinib or NVP-BGJ398 when compared to their FGFR2 wild-type counterparts (p=0.073 and p=0.021, respectively). Both agents inhibited FGFR2 signaling, induced cell cycle arrest and significantly increased apoptosis in FGFR2 mutant lines. In vitro, dovitinib and NVP-BGJ398 were both potent at inhibiting cell growth of FGFR2 mutant EC cells but the activity of dovitinib was less restricted to FGFR2 mutant lines when compared to NVP-BGJ398. In vivo, dovitinib and NVP-BGJ398 significantly inhibited the growth of FGFR2 mutated EC xenograft models. In addition, dovitinib showed significant antitumor activity in FGFR2 wild-type EC xenograft models including complete tumor regressions in a long term in vivo study. Dovitinib and NVP-BGJ398 warrant further clinical evaluation in patients with FGFR2 mutated EC. Dovitinib may have antitumor activity in EC beyond FGFR2 mutated cases and may permit greater flexibility in patient selection.

PMID: 23443805 [PubMed - as supplied by publisher]

c-met inhibitors zm-447439 rad001

Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-?/BMP signaling and orphan nuclear receptors.

Related Articles

Drosophila motor neuron retraction during metamorphosis is mediated by inputs from TGF-?/BMP signaling and orphan nuclear receptors.

PLoS One. 2012;7(7):e40255

Authors: Boulanger A, Farge M, Ramanoudjame C, Wharton K, Dura JM

Abstract
Larval motor neurons remodel during Drosophila neuro-muscular junction dismantling at metamorphosis. In this study, we describe the motor neuron retraction as opposed to degeneration based on the early disappearance of ?-Spectrin and the continuing presence of Tubulin. By blocking cell dynamics with a dominant-negative form of Dynamin, we show that phagocytes have a key role in this process. Importantly, we show the presence of peripheral glial cells close to the neuro-muscular junction that retracts before the motor neuron. We show also that in muscle, expression of EcR-B1 encoding the steroid hormone receptor required for postsynaptic dismantling, is under the control of the ftz-f1/Hr39 orphan nuclear receptor pathway but not the TGF-? signaling pathway. In the motor neuron, activation of EcR-B1 expression by the two parallel pathways (TGF-? signaling and nuclear receptor) triggers axon retraction. We propose that a signal from a TGF-? family ligand is produced by the dismantling muscle (postsynapse compartment) and received by the motor neuron (presynaptic compartment) resulting in motor neuron retraction. The requirement of the two pathways in the motor neuron provides a molecular explanation for the instructive role of the postsynapse degradation on motor neuron retraction. This mechanism insures the temporality of the two processes and prevents motor neuron pruning before postsynaptic degradation.

PMID: 22792255 [PubMed - indexed for MEDLINE]

rad001 ecdysone chir-258

DNA-PK phosphorylation of IGFBP-3 is required to Prevent Apoptosis in Retinal Endothelial cells cultured in High Glucose.

DNA-PK phosphorylation of IGFBP-3 is required to Prevent Apoptosis in Retinal Endothelial cells cultured in High Glucose.

Invest Ophthalmol Vis Sci. 2013 Apr 4;

Authors: Zhang Q, Steinle JJ

Abstract
PURPOSE: The goal of this study was to determine whether Compound 49b stimulates insulin-like growth factor binding protein-3 (IGFBP-3) activation in retinal endothelial cells (REC) through DNA-dependent protein kinase (DNA-PK). METHODS: REC were grown in normal glucose (5 mM) or high glucose medium (25 mM). Some cells were transfected with protein kinase A (PKA) siRNA, following treatment with 50 nM Compound 49b, a novel ?-adrenergic receptor agonist. Cell proteins were extracted and analyzed for DNA-PK expression by Western blotting. Additional cells were treated with or without NU7441 (a specific DNA-PK inhibitor) prior to Compound 49b treatment. Cell lysates were processed for IGFBP-3 ELISA analyses and Western blotting to measure casein kinase 2 (CK2). Immunoprecipitation for total and phospho-IGFBP-3 and cell death measurements were done after transfection with the S156A IGFBP-3 mutation (key phosphorylation site involved in DNA-PK) plasmid DNA. RESULTS: Compound 49b required DNA-PK to activate IGFBP-3 in REC. IGFBP-3 activation was significantly reduced following treatment with either the DNA-PK inhibitor or following transfection with the IGFBP-3 S156A mutant plasmid (p < 0.05). Significant increases in cell death were also observed in cells transfected with the IGFBP-3 S156A mutant plasmid (p < 0.05). Casein kinase levels were not altered after treatment with NU7741 or Compound 49b. CONCLUSIONS: Our finding suggested Compound 49b induces DNA-PK levels through PKA activity. DNA-PK is required for Compound 49b-induced activation of IGFBP-3, leading to inhibition of REC cell death.

PMID: 23557743 [PubMed - as supplied by publisher]

zm-447439 rad001 ecdysone

EMD 1214063 and EMD 1204831 constitute a new class of potent and highly selective c-Met inhibitors.

EMD 1214063 and EMD 1204831 constitute a new class of potent and highly selective c-Met inhibitors.

Clin Cancer Res. 2013 Apr 3;

Authors: Bladt F, Faden B, Friese-Hamim M, Knuehl C, Wilm C, Frittschen C, Graedler U, Meyring M, Dorsch D, Jaehrling F, Pehl U, Stieber F, Schadt O, Blaukat A

Abstract
PURPOSE: The mesenchymal-epithelial transition factor (c-Met) receptor, also known as hepatocyte growth factor receptor (HGFR), controls morphogenesis, a process that is physiologically required for embryonic development and tissue repair. Aberrant c-Met activation is associated with a variety of human malignancies including cancers of the lung, kidney, stomach, liver, and brain. In this study, we investigated the properties of two novel compounds, developed to selectively inhibit the c-Met receptor in anti-tumor therapeutic interventions. EXPERIMENTAL DESIGN: The pharmacologic properties, c-Met inhibitory activity, and anti-tumor effects of EMD 1214063 and EMD 1204831 were investigated in vitro and in vivo, using human cancer cell lines and mouse xenograft models. RESULTS: EMD 1214063 and EMD 1204831 selectively suppressed the c-Met receptor tyrosine kinase activity. Their inhibitory activity was potent (inhibitory 50% concentration [IC50], 3 nM and 9 nM, respectively) and highly selective, when compared to their effect on a panel of 242 human kinases. Both EMD 1214063 and EMD 1204831 inhibited c-Met phosphorylation and downstream signaling in a dose-dependent fashion, but differed in the duration of their inhibitory activity. In murine xenograft models, both compounds induced regression of human tumors, regardless of whether c-Met activation was HGF dependent or independent. Both drugs were well tolerated and induced no substantial weight loss after >3 weeks of treatment. CONCLUSIONS: Our results indicate selective c-Met inhibition by EMD 1214063 and EMD 1204831 and strongly support clinical testing of these compounds in the context of molecularly targeted anti-cancer strategies.

PMID: 23553846 [PubMed - as supplied by publisher]

c-met inhibitors zm-447439 rad001

DNA-PK phosphorylation of IGFBP-3 is required to Prevent Apoptosis in Retinal Endothelial cells cultured in High Glucose.

DNA-PK phosphorylation of IGFBP-3 is required to Prevent Apoptosis in Retinal Endothelial cells cultured in High Glucose.

Invest Ophthalmol Vis Sci. 2013 Apr 4;

Authors: Zhang Q, Steinle JJ

Abstract
PURPOSE: The goal of this study was to determine whether Compound 49b stimulates insulin-like growth factor binding protein-3 (IGFBP-3) activation in retinal endothelial cells (REC) through DNA-dependent protein kinase (DNA-PK). METHODS: REC were grown in normal glucose (5 mM) or high glucose medium (25 mM). Some cells were transfected with protein kinase A (PKA) siRNA, following treatment with 50 nM Compound 49b, a novel ?-adrenergic receptor agonist. Cell proteins were extracted and analyzed for DNA-PK expression by Western blotting. Additional cells were treated with or without NU7441 (a specific DNA-PK inhibitor) prior to Compound 49b treatment. Cell lysates were processed for IGFBP-3 ELISA analyses and Western blotting to measure casein kinase 2 (CK2). Immunoprecipitation for total and phospho-IGFBP-3 and cell death measurements were done after transfection with the S156A IGFBP-3 mutation (key phosphorylation site involved in DNA-PK) plasmid DNA. RESULTS: Compound 49b required DNA-PK to activate IGFBP-3 in REC. IGFBP-3 activation was significantly reduced following treatment with either the DNA-PK inhibitor or following transfection with the IGFBP-3 S156A mutant plasmid (p < 0.05). Significant increases in cell death were also observed in cells transfected with the IGFBP-3 S156A mutant plasmid (p < 0.05). Casein kinase levels were not altered after treatment with NU7741 or Compound 49b. CONCLUSIONS: Our finding suggested Compound 49b induces DNA-PK levels through PKA activity. DNA-PK is required for Compound 49b-induced activation of IGFBP-3, leading to inhibition of REC cell death.

PMID: 23557743 [PubMed - as supplied by publisher]

zm-447439 rad001 ecdysone

Association of a MET genetic variant with autism-associated maternal autoantibodies to fetal brain proteins and cytokine expression.

Related Articles

Association of a MET genetic variant with autism-associated maternal autoantibodies to fetal brain proteins and cytokine expression.

Transl Psychiatry. 2011;1:e48

Authors: Heuer L, Braunschweig D, Ashwood P, Van de Water J, Campbell DB

Abstract
The contribution of peripheral immunity to autism spectrum disorders (ASDs) risk is debated and poorly understood. Some mothers of children with ASD have autoantibodies that react to fetal brain proteins, raising the possibility that a subset of ASD cases may be associated with a maternal antibody response during gestation. The mechanism by which the maternal immune system breaks tolerance has not been addressed. We hypothesized that the mechanism may involve decreased expression of the MET receptor tyrosine kinase, an ASD risk gene that also serves as a key negative regulator of immune responsiveness. In a sample of 365 mothers, including 202 mothers of children with ASD, the functional MET promoter variant rs1858830 C allele was strongly associated with the presence of an ASD-specific 37+73-kDa band pattern of maternal autoantibodies to fetal brain proteins (P=0.003). To determine the mechanism of this genetic association, we measured MET protein and cytokine production in freshly prepared peripheral blood mononuclear cells from 76 mothers of ASD and typically developing children. The MET rs1858830 C allele was significantly associated with MET protein expression (P=0.025). Moreover, decreased expression of the regulatory cytokine IL-10 was associated with both the MET gene C allele (P=0.001) and reduced MET protein levels (P=0.002). These results indicate genetic distinction among mothers who produce ASD-associated antibodies to fetal brain proteins, and suggest a potential mechanism for how a genetically determined decrease in MET protein production may lead to a reduction in immune regulation.

PMID: 22833194 [PubMed - indexed for MEDLINE]

ecdysone chir-258 dovitinib

In Vivo c-Met Pathway Inhibition Depletes Human Glioma Xenografts of Tumor-Propagating Stem-Like Cells.

In Vivo c-Met Pathway Inhibition Depletes Human Glioma Xenografts of Tumor-Propagating Stem-Like Cells.

Transl Oncol. 2013 Apr;6(2):104-11

Authors: Rath P, Lal B, Ajala O, Li Y, Xia S, Kim J, Laterra J

Abstract
Solid malignancies contain sphere-forming stem-like cells that are particularly efficient in propagating tumors. Identifying agents that target these cells will advance the development of more effective therapies. Recent converging evidence shows that c-Met expression marks tumor-initiating stem-like cells and that c-Met signaling drives human glioblastoma multiforme (GBM) cell stemness in vitro. However, the degree to which tumor-propagating stem-like cells depend on c-Met signaling in histologically complex cancers remains unknown. We examined the effects of in vivo c-Met pathway inhibitor therapy on tumor-propagating stem-like cells in human GBM xenografts. Animals bearing pre-established tumor xenografts expressing activated c-Met were treated with either neutralizing anti- hepatocyte growth factor (HGF) monoclonal antibody L2G7 or with the c-Met kinase inhibitor PF2341066 (Crizotinib). c-Met pathway inhibition inhibited tumor growth, depleted tumors of sphere-forming cells, and inhibited tumor expression of stem cell markers CD133, Sox2, Nanog, and Musashi. Withdrawing c-Met pathway inhibitor therapy resulted in a substantial rebound in stem cell marker expression concurrent with tumor recurrence. Cells derived from xenografts treated with anti-HGF in vivo were depleted of tumor-propagating potential as determined by in vivo serial dilution tumor-propagating assay. Furthermore, daughter xenografts that did form were 12-fold smaller than controls. These findings show that stem-like tumor-initiating cells are dynamically regulated by c-Met signaling in vivo and that c-Met pathway inhibitors can deplete tumors of their tumor-propagating stem-like cells.

PMID: 23556031 [PubMed - in process]

ecdysone chir-258 dovitinib

Monoterpenoid indole alkaloids from Alstonia yunnanensis and their cytotoxic and anti-inflammatory activities.

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Monoterpenoid indole alkaloids from Alstonia yunnanensis and their cytotoxic and anti-inflammatory activities.

Molecules. 2012;17(11):13631-41

Authors: Cao P, Liang Y, Gao X, Li XM, Song ZQ, Liang G

Abstract
The 80% ethanol extract of Alstonia yunnanensis afforded five new monoterpenoid indole alkaloids: 11-hydroxy-6,7-epoxy-8-oxo-vincadifformine (1), 14-chloro-15-hydroxy- vincadifformine (2), perakine N(4)-oxide (3), raucaffrinoline N(4)-oxide (4), and vinorine N(1),N(4)-dioxide (5), together with three known compounds: 11-methoxy-6,7-epoxy-8-oxo- vincadifformine (6), vinorine N(4)-oxide (7) and vinorine (8). The structures of the isolated compounds were established based on 1D and 2D (1H-1H-COSY, HMQC, HMBC, and ROESY) NMR spectroscopy, in addition to high resolution mass spectrometry. The isolated compounds were tested in vitro for cytotoxic potential against seven tumor cell lines and anti-inflammatory activities. Compounds 3, 4 and 7 exhibited weak cytotoxicity against the tested cell lines and selective inhibition of Cox-2 (> 85%).

PMID: 23159924 [PubMed - indexed for MEDLINE]

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In Vivo c-Met Pathway Inhibition Depletes Human Glioma Xenografts of Tumor-Propagating Stem-Like Cells.

In Vivo c-Met Pathway Inhibition Depletes Human Glioma Xenografts of Tumor-Propagating Stem-Like Cells.

Transl Oncol. 2013 Apr;6(2):104-11

Authors: Rath P, Lal B, Ajala O, Li Y, Xia S, Kim J, Laterra J

Abstract
Solid malignancies contain sphere-forming stem-like cells that are particularly efficient in propagating tumors. Identifying agents that target these cells will advance the development of more effective therapies. Recent converging evidence shows that c-Met expression marks tumor-initiating stem-like cells and that c-Met signaling drives human glioblastoma multiforme (GBM) cell stemness in vitro. However, the degree to which tumor-propagating stem-like cells depend on c-Met signaling in histologically complex cancers remains unknown. We examined the effects of in vivo c-Met pathway inhibitor therapy on tumor-propagating stem-like cells in human GBM xenografts. Animals bearing pre-established tumor xenografts expressing activated c-Met were treated with either neutralizing anti- hepatocyte growth factor (HGF) monoclonal antibody L2G7 or with the c-Met kinase inhibitor PF2341066 (Crizotinib). c-Met pathway inhibition inhibited tumor growth, depleted tumors of sphere-forming cells, and inhibited tumor expression of stem cell markers CD133, Sox2, Nanog, and Musashi. Withdrawing c-Met pathway inhibitor therapy resulted in a substantial rebound in stem cell marker expression concurrent with tumor recurrence. Cells derived from xenografts treated with anti-HGF in vivo were depleted of tumor-propagating potential as determined by in vivo serial dilution tumor-propagating assay. Furthermore, daughter xenografts that did form were 12-fold smaller than controls. These findings show that stem-like tumor-initiating cells are dynamically regulated by c-Met signaling in vivo and that c-Met pathway inhibitors can deplete tumors of their tumor-propagating stem-like cells.

PMID: 23556031 [PubMed - in process]

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