Objective: To explore the effect of the AXL expression on the chemosensitivity of prostate cancer PC-3 and DU145 cells to docetaxel and possible mechanisms. METHODS: Using Western blot, we examined the expressions of the AXL protein, p-AXL and Gas6 in the docetaxel-resistant PC-3 (PC-3-DR) and DU145 (DU145-DR) cells stimulated with gradually increased concentrations of docetaxel. We transfected the PC-3 and DU145 cells with negative NC ShRNA and AXL-ShRNA, respectively, which were confirmed to be effective, detected the proliferation, apoptosis and cycle distribution of the cells by CCK8, MTT and flow cytometry after treated with the AXL-inhibitor MP470 and/or docetaxel, and determined the expression of the ABCB1 protein in the PC-3-DR and DU145-DR cells after intervention with the AXL-inhibitor R428 and/or docetaxel. RESULTS: The expression of the AXL protein in the PC-3 and DU145 cells was significantly increased after docetaxel treatment (P <0.05). The expressions AXL and p-AXL were remarkably higher (P <0.05) while that of Gas6 markedly lower (P <0.05) in the PC-3 and DU145 than in the PC-3-DR and DU145-DR cells. The inhibitory effect of docetaxel on the proliferation and its enhancing effect on the apoptosis of the PC-3 and DU145 cells were significantly decreased at 48 hours after AXL transfection (P <0.05). MP470 obviously suppressed the growth and promoted the apoptosis of the PC-3-DR and DU145-DR cells, with a higher percentage of the cells in the G2/M phase when combined with docetaxel than used alone (P <0.05). R428 markedly reduced the expression of ABCB1 in the PC-3-DR and DU145-DR cells, even more significantly in combination with docetaxel than used alone (P <0.05). CONCLUSIONS The elevated expression of AXL enhances the docetaxel-resistance of PC-3 and DU145 prostate cancer cells and AXL intervention improves their chemosensitivity to docetaxel, which may be associated with the increased cell apoptosis in the G2/M phase and decreased expression of ABCB1.
ATP Binding Cassette Transporter, Subfamily B, Member 1 ; metabolism ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Cell Count ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Docetaxel ; Drug Resistance, Neoplasm ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Male ; Prostatic Neoplasms ; drug therapy ; metabolism ; pathology ; Proto-Oncogene Proteins ; drug effects ; genetics ; metabolism ; Pyrimidines ; pharmacology ; RNA, Small Interfering ; Receptor Protein-Tyrosine Kinases ; drug effects ; genetics ; metabolism ; Taxoids ; pharmacology

Objective: To study the effects of muskolibanum combination on the proliferation and differentiation of prostate stem cells. METHODS: We cultured prostate epithelial cells and urogenital sinus mesenchymal (UGSM) cells from 7－10 d old C57BL/6 mice and 16－18 d old pregnant C57BL/6 mice, transplanted the mixed suspension of the two types of cells under the kidney envelope of SCIDCB.17 male mice, and harvested the transplants 30 days later. We randomly divided the SCIDCB.17 mice into four groups to be treated intragastrically with musk (n = 8), olibanum (n = 8), musk+olibanum (n = 7), and normal saline (blank control, n = 8)) respectively, all for 14 days. Then we collected the kidney tissue for observation of the morphology of the glandular tubes and differentiation of different subsets of stem cells by HE staining and determination of the expressions and distribution of P63, CD133, CD117 and Sca1 by immunohistochemistry and Western blot. RESULTS: A system was successfully established for the isolation and mixed culture of Sca1 Lin+ CD49f+ (LSC) cells of prostate stem cells and UGSM cells of the mouse embryonic prostate. Immunohistochemistry showed positive expressions of P63, CD133, Sca1, and CD117 in the prostatic acinar epithelia and proved the presence of prostatic acinar epithelial structure in the transplants. Compared with the blank control group, the expressions of CD133, Sca1 and CD117 were significantly increased in the musk, olibanum, and musk+olibanum groups (P< 0.05), higher in the musk+olibanum than in the musk or olibanum group (P< 0.05), and their protein expressions were even more elevated in the musk+olibanum group (P< 0.01), with statistically significant difference from the olibanum group (P< 0.05). CONCLUSIONS The combination of musk and olibanum can improve the proliferation and differentiation of prostate stem cells.
Animals ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Drug Therapy, Combination ; Epithelial Cells ; cytology ; drug effects ; Fatty Acids, Monounsaturated ; pharmacology ; Female ; Frankincense ; pharmacology ; Male ; Mesenchymal Stem Cells ; cytology ; drug effects ; Mice ; Mice, Inbred C57BL ; Mice, SCID ; Pregnancy ; Prostate ; cytology ; Random Allocation ; Receptor Protein-Tyrosine Kinases ; Receptors, Cholinergic ; Stem Cells ; cytology ; drug effects

Axl encodes the tyrosine-protein kinase receptor, participating in the proliferation and migration of many cells. This study examined the role of Axl in functions of endothelial progenitor cells (EPCs). Axl was detected by RT-PCR and Western blotting in both placentas and EPCs from normal pregnancy and preeclampsia patients. The Axl inhibitor, BMS777-607, was used to inhibit the Axl signalling pathway in EPCs. Cell proliferation, differentiation, migration and adhesion were measured by CCK-8 assay, cell differentiation assay, Transwell assay, and cell adhesion assay, respectively. Results showed the expression levels of Axl mRNA and protein were significantly higher in both placentas and EPCs from preeclampsia patients than from normal pregnancy (P<0.05). After treatment with BMS777-607, proliferation, differentiation, migration and adhesion capability of EPCs were all significantly decreased. Our study suggests Axl may play a role in the function of EPCs, thereby involving in the pathogenesis of preeclampsia.
Adult ; Aminopyridines ; pharmacology ; Blood Pressure ; Case-Control Studies ; Cell Adhesion ; drug effects ; Cell Differentiation ; drug effects ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Female ; Fetal Blood ; cytology ; enzymology ; Gene Expression Regulation ; Gestational Age ; Human Umbilical Vein Endothelial Cells ; drug effects ; enzymology ; pathology ; Humans ; Placenta ; metabolism ; physiopathology ; Pre-Eclampsia ; blood ; genetics ; physiopathology ; Pregnancy ; Primary Cell Culture ; Protein Kinase Inhibitors ; pharmacology ; Proto-Oncogene Proteins ; antagonists & inhibitors ; genetics ; metabolism ; Pyridones ; pharmacology ; RNA, Messenger ; antagonists & inhibitors ; genetics ; metabolism ; Receptor Protein-Tyrosine Kinases ; antagonists & inhibitors ; genetics ; metabolism ; Stem Cells ; drug effects ; enzymology ; pathology

OBJECTIVEThis study aims to evaluate the efficacy and safety of crizotinib for advanced ALK-positive non-small cell lung cancer (NSCLC) patients.

METHODSTwenty-eight patients with advanced ALK-positive NSCLC were given orally crizotinib 250 mg b. i.d., and were followed up to evaluate the therapeutic efficacy and safety.

RESULTSAmong the 28 patients, the objective response rate (ORR) was 71.4% (20/28) and disease control rate (DCR) was 92.9% (26/28). Three patients achieved complete response. Seventeen patients had partial response. The most common drug-related adverse events were mild flickering vision and gastrointestinal reaction. Eleven patients experienced flickering vision. Nine patients had nausea and vomiting. Eight patients had diarrhea. They were all reversible and of grade I or II. Only one patient had grade III myelosuppression. Among the 28 patients, 16 cases were disease-free and 12 cases had progressive disease, with a progression-free survival of 8.2 months.

CONCLUSIONSCrizotinib is effective and tolerable in the treatment of advanced ALK-positive NSLCC. However, its long-term treatment efficacy requires to be further studied.

Antineoplastic Agents ; adverse effects ; therapeutic use ; Carcinoma, Non-Small-Cell Lung ; drug therapy ; enzymology ; pathology ; Diarrhea ; chemically induced ; Disease-Free Survival ; Humans ; Lung Neoplasms ; drug therapy ; enzymology ; pathology ; Nausea ; chemically induced ; Protein Kinase Inhibitors ; adverse effects ; therapeutic use ; Pyrazoles ; adverse effects ; therapeutic use ; Pyridines ; adverse effects ; therapeutic use ; Receptor Protein-Tyrosine Kinases ; Vomiting ; chemically induced

OBJECTIVETo study the proto-oncogene RON mediated aggression of Raji cells and the inhibitory effects by monoclonal antibody Zt/f2 (2f2).

METHODSThe effects of RON ligand macrophage stimulating protein (MSP) (2.0 nmol/L) and inhibitory Zt/f2 (2F2) (2.0 nmol/L) antibody on proliferation of RON positive Raji cells after treatment for 24 and72 hours were detected by MTT method, colony formation units (CFU) of Raji cells by methylcellulose semi solid culture, Raji cells apoptosis and cell cycle analysis by AnnexinV/PI double staining, expression of RON, apoptosis-related proteins, and cyclins by Western blot.

RESULTS(1)Compared with the cell viability (1.0) and counts of CFU (103.6±7.0) in control group, Raji cells after MSP treatment had better viability (1.35±0.20) and CFU counts (133.7±10.4) (P<0.05), but worse viability (0.68±0.11) and CFU counts (66.3±6.1) after Zt/f2 (2F2) treatment (P<0.05). (2)Percentage of Raji cells apoptosis after Zt/f2 (2F2) antibody treatment (12.16±2.33)% was significantly increased than the control (2.89±1.03)% (P<0.05). The percentage of Raji cells arrested in G0/G1 phase was increased after Zt/f2 (2F2) antibody treatment as compared to the control [ (54.96 ±3.70)% vs (39.10±2.30)%, (P<0.05) ]. (3) High-level of RON phosphorylation and β-catenin expression activated by MSP could be inhibited significantly by Zt/f2 (2F2), which also up-regulated the expression of caspase-3, caspase-8, caspase-9 and PARP and down-regulated anti-apoptotic MCL-1 gene and inhibitor of apoptosis protein XIAP expression, accompanied with G1 phase protein changes accordingly.

CONCLUSIONMSP could aggravate Raji cells proliferation. Inversely, Zt/f2 (2F2) could inhibit proliferation and induce apoptosis by inhibition of RON phosphorylation and up-regulation of apoptosis related proteins.

Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Proto-Oncogenes ; Receptor Protein-Tyrosine Kinases ; metabolism

OBJECTIVETo explore correlation of seven apoptosis-related proteins (Hsp90a, p53, MDM2, Bcl-2, Bax, Cytochrome C, and Cleaved caspase3) with clinical outcomes of ALK+ anaplastic large-cell lymphoma (ALCL).

METHODSUsing immunohistochemistry and immunofluorescence double staining methods, the expressions of these seven apoptosis-associated proteins were studied to clarify their relationship with clinical outcomes of 36 ALK+ and 25 ALK-systemic ALCL patients enrolled between 1996 and 2006. The relationship of these apoptosis-regulating proteins with NPM-ALK status was also evaluated with the tyrosine inhibitor herbimycin A (HA) in vitro by immunocytochemistry, Western blotting and flow cytometric assays.

RESULTSThe presence of Hsp90α-, MDM2-, Bax-, Cytochrome C, and Cleaved caspase3-positive tumor cells was found significantly different in ALK+ and ALK-ALCLs, which was correlated with highly favorable clinical outcome. The Bcl-2- and p53-positive tumor cells were found in groups of patients with unfavorable prognosis. Inhibition of NPM-ALK by HA could reactivate the p53 protein and subsequent apoptosis-related proteins and therefore induced apoptosis in ALK+ ALCL cells.

CONCLUSIONOur results suggest that these seven proteins might be involved in apoptosis regulation and associated with clinical outcome of ALK+ systemic ALCLs. We also reveal a dynamic chain relation that NPM-ALK regulates p53 expression and subsequent apoptosis cascade in ALK+ ALCLs.

Adolescent ; Adult ; Aged ; Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; metabolism ; Benzoquinones ; pharmacology ; Biomarkers, Tumor ; metabolism ; Blotting, Western ; Cell Culture Techniques ; Cell Survival ; drug effects ; Child ; Child, Preschool ; Disease-Free Survival ; Enzyme Inhibitors ; pharmacology ; Female ; Flow Cytometry ; Humans ; Immunohistochemistry ; Kaplan-Meier Estimate ; Lactams, Macrocyclic ; pharmacology ; Lymphoma, Large-Cell, Anaplastic ; enzymology ; metabolism ; pathology ; Male ; Microscopy, Fluorescence ; Middle Aged ; Neoplasm Staging ; Prognosis ; Protein-Tyrosine Kinases ; metabolism ; Receptor Protein-Tyrosine Kinases ; antagonists & inhibitors ; metabolism ; Retrospective Studies ; Rifabutin ; analogs & derivatives ; Young Adult

Silibinin, from milk thistle (Silybum marianum), is a flavonolignan with anti-oxidative and anti-inflammatory properties. It has been therapeutically used for the treatment of hepatic diseases in China, Germany and Japan. Recently, increasing evidences prove that silibinin is also a potent antitumor agent, and the major anti-tumor mechanism for silibinin is the prominent inhibition of the activities of receptor tyrosine kinases (RTKs) and their downstream signal molecules in a variety of tumor cell lines, such as epidermal growth factor receptor 1 (EGFR) and insulin-like growth factor 1 receptor (IGF-1R) signaling pathways. Meanwhile, the evidences that silibinin selectively scavenges hydroxyl free radical (*OH) and specifically inhibits the action of nuclear factor kappaB (NF-kappaB) provide more complicated explanations for its antioxidant and anti-inflammatory effects. Some new findings such as that silibinin attenuating the cognitive deficits induced by amyloid beta protein (Abeta) peptide through its antioxidative and anti-inflammatory properties is valuable to broad the medical prospect of silibinin. In this review, we discuss the molecular pharmacological mechanisms of silibinin, focusing on its inhibition of tyrosine kinases, actions of antioxidation, free radical scavenging, immunoregulation and anti-inflammation.
Amyloid beta-Peptides ; metabolism ; Animals ; Anti-Inflammatory Agents ; pharmacology ; Antineoplastic Agents, Phytogenic ; pharmacology ; Antioxidants ; pharmacology ; Enzyme Activation ; Free Radical Scavengers ; pharmacology ; Humans ; Milk Thistle ; chemistry ; Molecular Structure ; NF-kappa B ; metabolism ; Protein-Tyrosine Kinases ; metabolism ; Reactive Oxygen Species ; metabolism ; Receptor Protein-Tyrosine Kinases ; metabolism ; Receptor, Epidermal Growth Factor ; metabolism ; Receptor, IGF Type 1 ; metabolism ; Signal Transduction ; drug effects ; Silymarin ; chemistry ; isolation & purification ; pharmacology

OBJECTIVETo investigate the influence of Qingkailing (QKL) on learning and memory abilities, global neurotransmitter and phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) signaling pathway of senescence accelerated mouse-prone/8 (SAMP8) mice with Alzheimer's dementia (AD).

METHODSSAMP mice were modeled and divided into the model group, the QKL group and the doneppezil hydrochloride group, all treated for 90 days. And a control group was set up with senescence accelerated mouse-resistance/1 (SAMR1) mice. Morris water maze was used to test the learning and memory abilities of mice; contents of acetylcholine (Ach) and monoamine neurotransmitters in brain were measured by HPLC; levels of Grb2-associated binder-1 (Gab1), AKT and phospho-serine/threonine protein kinase B (PAKT473) were evaluated by Western-blot.

RESULTSCompared with the control group, in the model group, the average escape latency detected by hidden platform trial and reverse trial on the 3rd day was higher (P < 0.01); levels of Ach, 5-hydroxytryptamine (5-HT) and Gab1 were lower (P < 0.01, P < 0.05 and P < 0.01), respectively. As compared with the model group, the escape latency (within the 2nd to 5th day) decreased (P < 0.01), levels of Ach and 5-HT increased (P < 0.05), and Gab1 protein expression increased (P < 0.01) in the QKL treated group after treatment, in addition, the level of phosphorylated AKT protein significantly increased (P < 0.05).

CONCLUSIONQKL could improve the learning and memory ability of AD model mice, which is probably related to its function in increasing cerebral Ach, 5-HT and activating PI3K/AKT pathway.

Alzheimer Disease ; drug therapy ; physiopathology ; Animals ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Female ; Learning ; drug effects ; Male ; Memory ; drug effects ; Mice ; Mice, Mutant Strains ; Phosphatidylinositol 3-Kinases ; metabolism ; Phytotherapy ; Proto-Oncogene Proteins ; metabolism ; Receptor Protein-Tyrosine Kinases ; metabolism ; Signal Transduction ; drug effects

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

5-Aryl-4-cyano-1H-1, 2, 3-triazoles bearing a variety of substituting groups on 5-phenyl were synthesized. Their structures were established by MS, IR and 1H NMR spectra. The crystal structures of compounds 3f and 3m were determined by X-ray diffraction analysis. The active H of the triazole was on 1-N from the crystal structures. The compounds, designed as HER2 tyrosine kinase inhibitors, were screened for bioactivity of growth-inhibition of breast cancer MDA-MB-453 cells. The lowest IC50 value of inhibiting HER2 tyrosine kinase phosphorylation in breast cancer cells is 6.6 micromol x L(-1). The inhibiting-growth of breast cancer cells was enhanced from electron-drawing groups joining 5-phenyl on the triazole.
Breast Neoplasms ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Crystallization ; Crystallography, X-Ray ; Female ; Humans ; Phosphorylation ; Protein-Tyrosine Kinases ; antagonists & inhibitors ; metabolism ; Receptor, ErbB-2 ; antagonists & inhibitors ; metabolism ; Triazoles ; chemical synthesis ; chemistry ; pharmacology