Genistein is a high specific and noncompetitive inhibitor of epidermal growth factor receptor tyramine kinase domain (EGFR-TK). In the paper, a molecular docking between genistein and EGFR-TK was studied to explore the mechanism of their interaction and antitumor mechanism of genistein by AUTODOCK 3.05 program. The results indicated that genistein located in the active cavity of EGFR-TK by high affinity (deltaG = -31.2 kJ/mol), and genistein inhibited EGFR-TK by interfering with forming of Lys721/Glu738 ion pair. The inhibition belonged to noncompetitive interaction, in which hydrophobic force and hydrogen bond played key roles.
Catalytic Domain ; Genistein ; metabolism ; pharmacology ; Models, Molecular ; Receptor, Epidermal Growth Factor ; antagonists & inhibitors ; metabolism

OBJECTIVETo build 3D-pharmacophore model of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors using distance comparisons method and design novel EGFR inhibitors.

METHODSThirteen typical EGFR inhibitors were selected, and their biologically active conformations were obtained by using DOCK5.0 program, then 3D-pharmacophore model of EGFR inhibitors was built using distance comparisons method.

RESULTSValidation of the 3D-pharmacophore model was carried out and novel structures with potential inhibitory activity were selected by means of 3D-searching and docking method.

CONCLUSIONThis method can improve hit rate of lead compounds discovery and can be used to design novel EGFR inhibitors.

Drug Design ; Enzyme Inhibitors ; Epidermal Growth Factor ; metabolism ; Models, Chemical ; Protein-Tyrosine Kinases ; antagonists & inhibitors ; chemistry ; pharmacology ; Receptor, Epidermal Growth Factor ; antagonists & inhibitors ; chemistry ; Structure-Activity Relationship

OBJECTIVETo establish an in vitro homogeneous time-resolved fluorescence immunoassay method for high throughput screening of protein tyrosine kinase (PTK) inhibitors.

METHODSSpecific fluorescence signals at 670 and 612 nm were measured by multifunctional microplate reader when the fluorescence was emitted through a resonance energy transfer between fluorescent materials (EuK and XL-665). The inhibitory activity of Sunitinib, a standard PTK inhibitor, on vascular endothelia growth factor receptor 2 (VEGFR-2) kinase activity was investigated.

RESULTSA homogeneous time-resolved fluorescence immunoassay was established for high throughput screening of PTK inhibitor. In this system, the concentrations of VEGFR-2, adenosine triphosphate (ATP) and poly-peptide substrate were 5 ng/microl, 100 micromol/L and 1 micromol/L, respectively. Sunitinib inhibited VEGFR-2 kinase activity with an IC50 value of 86.7 nmol/L, which was close to the values tested using other methods.

CONCLUSIONThe homogeneous time-resolved fluorescence immunoassay we established can be easily used for high throughput screening of PTK inhibitors.

Fluoroimmunoassay ; methods ; High-Throughput Screening Assays ; methods ; Indoles ; pharmacology ; Peptides ; metabolism ; Phosphorylation ; drug effects ; Protein Kinase Inhibitors ; pharmacology ; Protein-Tyrosine Kinases ; antagonists & inhibitors ; metabolism ; Pyrroles ; pharmacology ; Time Factors ; Vascular Endothelial Growth Factor Receptor-2 ; antagonists & inhibitors ; metabolism

This study was aimed to investigate the influence and significance of celecoxib (specific inhibitor of cyclooxygenase-2) on mRNA expressions of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (b-FGF), transforming growth factor β (TGF-β) in acute leukemia cells. The expressions of VEGF, b-FGF, TGF-β mRNA were measured by RT-PCR in acute leukemia cells treated with celecoxib (80 µmol/L, for 48 h) or with PBS. The results showed that the obvious expressions of VEGF, b-FGF, TGF-β mRNA were observed in acute leukemia cells. By using Pearson correlation analysis, there was positive correlation between VEGF mRNA and b-FGF mRNA expressions (r = 0.559, P = 0.001), and negative correlation between VEGF and TGF-β mRNA expressions (r = -0.4, P = 0.029). Expression levels of VEGF, b-FGF, TGF-β mRNA in experimental group were lower than that in control group (P < 0.01). It is concluded that COX-2 inhibitor celecoxib can inhabit vascular endothelial growth through down-regulating the mRNA expression of VEGF, b-FGF and TGF-β in acute leukemia cells. COX-2 inhibitor may offer supplemental effect for treating acute leukemia.
Adult ; Celecoxib ; Cyclooxygenase 2 Inhibitors ; pharmacology ; Female ; Fibroblast Growth Factor 2 ; metabolism ; Humans ; Leukemia ; drug therapy ; metabolism ; Male ; Pyrazoles ; pharmacology ; RNA, Messenger ; genetics ; Signal Transduction ; Sulfonamides ; pharmacology ; Transforming Growth Factor beta ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism

This paper sums up some studies in the last decade regarding the inhibitory effects of traditional Chinese herbs on growth factor induced proliferation of vascular smooth muscle cell (VSMC) via directly measuring the mRNA expression of its growth factors and the related receptors by electron microscope, immunohistochemistry, blot and hybridization in situ.
Cell Proliferation ; drug effects ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Fibroblast Growth Factor 2 ; antagonists & inhibitors ; Humans ; Muscle, Smooth, Vascular ; cytology ; metabolism ; Platelet-Derived Growth Factor ; antagonists & inhibitors

BACKGROUNDIt has been reported that there is a significant difference in the local tissue concentration of transforming growth factor (TGF)-β1 between chronic rhinosinusitis without nasal polyps (CRSsNP) and chronic rhinosinusitis without nasal polyps (CRSwNP) patients. TGF-β has been reported to play an important role in regulating epithelial cell repair in lower airway remodeling and may be a critical factor involved in the remodeling process of chronic rhinosinusitis (CRS).

METHODSEthmoidal mucosal samples collected from CRS and healthy control patients were analyzed for TGF-β1, TGF-β receptor I, TGF-β receptor II, Smad3, phospho-Smad3, Smad7, and Smad anchor for receptor activation by Western blotting analysis. The proliferation of sinonasal epithelial cells at baseline and after TGF-β1 and/or EGF stimulation was evaluated by the MTT assay.

RESULTSIn CRSsNP, TGF-β1, TGF-β receptor I, TGF-β receptor II, and Smad3 protein levels were significantly higher than controls. In CRSwNP, TGF-β1, Smad3, and pSmad3 protein levels were significantly lower than controls. Smad7 protein was significantly higher in CRS than controls. In vitro experiments demonstrated that the baseline proliferation levels of sinonasal epithelial cells were lower in CRS than controls.

CONCLUSIONSCRSwNP is characterized by a lower level of TGF-signaling compared with the control. In CRSsNP, although the upstream signaling of TGF-β was enhanced, the high Smad7 protein expression may restrain the downstream signaling components (e.g., pSmad3) and the TGF-β antiproliferative effect on sinonasal epithelium. The difference in the local tissue concentration of TGF-β1 between CRSsNP and CRSwNP patients did not result in significant differences in epithelial proliferation.

Adult ; Aged ; Benzamides ; pharmacology ; Cells, Cultured ; Dioxoles ; pharmacology ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Male ; Middle Aged ; Protein-Serine-Threonine Kinases ; antagonists & inhibitors ; metabolism ; Receptors, Transforming Growth Factor beta ; antagonists & inhibitors ; metabolism ; Serine Endopeptidases ; metabolism ; Signal Transduction ; drug effects ; Sinusitis ; metabolism ; Smad3 Protein ; metabolism ; Smad7 Protein ; metabolism ; Transforming Growth Factor beta ; metabolism ; Transforming Growth Factor beta1 ; antagonists & inhibitors ; metabolism ; Young Adult

Angiogenesis Inhibitors ; pharmacology ; Angiostatins ; pharmacology ; Animals ; Antigens, CD ; metabolism ; Cadherins ; metabolism ; Humans ; Matrix Metalloproteinase Inhibitors ; Matrix Metalloproteinases ; metabolism ; Microvessels ; metabolism ; Neoplasms ; blood supply ; pathology ; Neovascularization, Pathologic ; metabolism ; pathology ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVETo screen for the inhibitor of vascular endothelial growth factor (VEGF) 165 from random peptide library.

METHODSPositive phage clones were rescued after two rounds of panning and competitive elution. Its affinity activity to KDR was monitored through ELISA, immunohistochemical method, Chicken CAM assay and MTT.

RESULTSFive specific binding positive target molecule phage clones were obtained which were able to bind to cells whose surface had high KDR, among which, clone 3 and 13 could effectively block the vascularization of the chorioallantoic membrane of chick embryo, but they were not inhibitive on the proliferation of high KDR expression cells.

CONCLUSIONThe peptides, being the inhibitors of VEGF, may be useful in the treatment of cancers.

Animals ; Binding Sites ; Endothelial Growth Factors ; antagonists & inhibitors ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Lymphokines ; antagonists & inhibitors ; metabolism ; Peptide Library ; Peptides ; pharmacology ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors

Lysophosphatidic acid (LPA) is a bioactive phospholipids and involves in various cellular events, including tumor cell migration. In the present study, we investigated LPA receptor and its transactivation to EGFR for cyclooxygenase-2 (COX-2) expression and cell migration in CAOV-3 ovarian cancer cells. LPA induced COX-2 expression in a dose-dependent manner, and pretreatment of the cells with pharmacological inhibitors of Gi (pertussis toxin), Src (PP2), EGF receptor (EGFR) (AG1478), ERK (PD98059) significantly inhibited LPA- induced COX-2 expression. Consistent to these results, transfection of the cells with selective Src siRNA attenuated COX-2 expression by LPA. LPA stimulated CAOV-3 cell migration that was abrogated by pharmacological inhibitors and antibody of EP2. Higher expression of LPA2 mRNA was observed in CAOV-3 cells, and transfection of the cells with a selective LPA2 siRNA significantly inhibited LPA-induced activation of EGFR and ERK, as well as COX-2 expression. Importantly, LPA2 siRNA also blocked LPA-induced ovarian cancer cell migration. Collectively, our results clearly show the significance of LPA2 and Gi/Src pathway for LPA-induced COX-2 expression and cell migration that could be a promising drug target for ovarian cancer cell metastasis.
Butadienes/pharmacology ; Cell Line, Tumor ; Cell Movement/drug effects/*physiology ; Cyclooxygenase 2/*biosynthesis ; Extracellular Signal-Regulated MAP Kinases/antagonists & inhibitors/metabolism ; Female ; Flavonoids/pharmacology ; GTP-Binding Protein alpha Subunits, Gi-Go/antagonists & inhibitors/*metabolism ; Humans ; Lysophospholipids/pharmacology ; Nitriles/pharmacology ; Ovarian Neoplasms/metabolism/*pathology ; Pertussis Toxin/pharmacology ; Protein-Tyrosine Kinases/antagonists & inhibitors/*metabolism ; Proto-Oncogene Proteins/antagonists & inhibitors/*metabolism ; Pyrimidines/pharmacology ; Receptor, Epidermal Growth Factor/antagonists & inhibitors/metabolism ; Receptors, Lysophosphatidic Acid/*metabolism ; Receptors, Prostaglandin E/metabolism ; Signal Transduction ; Transcriptional Activation ; Tyrphostins/pharmacology

OBJECTIVETo investigate the effects of pioglitazone on transforming growth factor beta1 (TGFbeta1) expression in ischemia/reperfusion injury myocardium of rats.

METHODSThirty SD rats were randomly divided into five groups (n = 6): ischemia/reperfusion group, pioglitazone 5 mg/(kg x d) group, pioglitazone 10 mg/(kg x d) group, pioglitazone 20 mg/(kg x d) group and pioglitazone 20 mg/(kg x d) + peroxisome proliferator-activated receptor gamma (PPARgamma) specific antagonist GW9662 group. Left anterior descending coronary artery of rats were ligated for 30 min and reperfused for 120 min to establish the model of ischemia/reperfusion in vivo. RT-PCR was performed to detect the expression of TGFbeta1 mRNA. Western blot was performed to detect the expression of TGFbeta1 protein.

RESULTSMyocardial apoptosis was significantly suppressed by pioglitazone. Pioglitazone upregulated TGFPbeta1 expression both in mRNA and protein level. GW9662 reversed the inhibition of myocardial apoptosis and the upregulation of TGFbeta1 expression by pioglitazone.

CONCLUSIONPioglitazone can inhibit the myocardial apoptosis induced by ischemia/reperfusion. Pioglitazone may protect the myocardium from ischemia/reperfusion via upregulation of TGFbeta1. This protection may be mediated by PPARgamma.

Anilides ; pharmacology ; Animals ; Apoptosis ; Male ; Myocardial Reperfusion Injury ; metabolism ; Myocardium ; metabolism ; PPAR gamma ; antagonists & inhibitors ; Rats ; Rats, Sprague-Dawley ; Thiazolidinediones ; pharmacology ; Transforming Growth Factor beta1 ; metabolism