Journal: PLoS ONE

Article Title: A Novel Requirement for Janus Kinases as Mediators of Drug Resistance Induced by Fibroblast Growth Factor-2 in Human Cancer Cells

doi: 10.1371/journal.pone.0019861

Figure Lengend Snippet: A. FGF-2 did not induce STAT1, STAT3, or STAT5 phosphorylation. U2OS cells were serum-starved overnight and then stimulated with FGF-2 for the indicated times. Cells treated with IFN-γ (500 IU/ml), IL-6 (200 ng/ml)/sIL-6R (250 ng/ml) and OSM (50 ng/ml) were used as positive controls for activation of STAT1, STAT3 and STAT5, respectively. Proteins were analyzed as before using antibodies against phosphorylated STAT1 (Tyr 701 ), phosphorylated STAT3 (Tyr 705 ), phosphorylated STAT5 (Tyr 694 ) and antibodies that recognize both phosphorylated and unphosphorylated proteins. B. FGF-2 induced the phosphorylation of TYK2 in U2OS cells. Cells were incubated in serum-free media and then treated with FGF-2 (10 ng/ml) for the indicated times. TYK2 was immunoprecipitated and western blotting analysis was performed using antibodies against phosphorylated tyrosines and total TYK2. TYK2 was used as a loading control. C. Total cell lysates used to immunoprecipitate TYK2 and from cells transfected with siTYK2 were separated on a 7.5% SDS-PAGE gel and analyzed by western blot. Membranes were probed for TYK2, pERK1/2-Thr 202/185 /Tyr 204/187 and total ERK1/2. β-actin was used as a loading control. ‘ – min.

Article Snippet: Antibodies against phosphorylated STAT1 (pSTAT1-Tyr 701 ), phosphorylated STAT3 (pSTAT3-Tyr 705 ), phosphorylated STAT5 (pSTAT5-Tyr 694 ), phosphorylated ERK1/2 (pERK1/2-Thr 202/185 /Tyr 204/187 ) and PARP were bought from Cell Signaling Technology.

Techniques: Phospho-proteomics, Activation Assay, Incubation, Immunoprecipitation, Western Blot, Control, Transfection, SDS Page

Journal: Journal of Cancer

Article Title: CSF2 Overexpression Is Associated with STAT5 Phosphorylation and Poor Prognosis in Patients with Urothelial Carcinoma

doi: 10.7150/jca.14281

Figure Lengend Snippet: Summary of two significantly and differentially expressed genes related to positive regulation of tyrosine phosphorylation of pSTAT5 in the published transcriptome of UBUC (GSE32894).

Article Snippet: The endogenous peroxidase was quenched by saline for 15 minutes and then incubated with primary monoclonal antibodies against CSF2 (1:100, Cat. No. ab77768, rabbit polyclonal, abcam, Cambridge, MA) and phosphor-STAT5 (pSTAT5, Tyr 694/Tyr 699) (1:50, Cat. No. sc-11761, goat polyclonal, Santa Cruz, CA) for an hour.

Techniques: Phospho-proteomics, Cell Differentiation, Expressing, Activity Assay, Binding Assay, Transduction, Migration, Protein Binding

Journal: Journal of pharmacokinetics and pharmacodynamics

Article Title: FLT3 and CDK4/6 inhibitors: Signaling mechanisms and tumor burden in subcutaneous and orthotopic mouse models of acute myeloid leukemia

doi: 10.1007/s10928-014-9393-x

Figure Lengend Snippet: Plasma PK- cellular signaling- tumor burden modeling scheme. AMG925, sorafenib and AC220 inhibit STAT5 phosphorylation via inhibiting FLT3ITD. AMG925 inhibits Rb phosphorylation by directly targeting CDK4/6, and sorafenib also influences the activity of cycD1·CDK4/6 via inhibitory effects on other receptors and/or kinases, including VEGFR, PDGFR-β, c-KIT and etc. The decreased pSTAT5 values not only promote apoptosis in tumor cells, but also hinder the proliferation of tumor cells. In the modeling analysis, PK parameters were fixed at values in Table 1 during parameter estimation of cellular signaling models, and cellular signaling model parameters were fixed at estimates in Table 2 (Subcutaneous tumor studies) or Table 4 (Orthotopic tumor studies) while estimating parameters of the tumor burden models.

Article Snippet: Lysates were prepared and analyzed for STAT5 Y694 phosphorylation and Rb S780 phosphorylation using MSD (Meso Scale Discovery) assays, which were then normalized to total STAT5 and p38 MAPK values to produce the values of phosphorylated STAT5 (pSTAT5) and of phosphorylated Rb (pRb).

Techniques: Clinical Proteomics, Phospho-proteomics, Activity Assay

Journal: Journal of pharmacokinetics and pharmacodynamics

Article Title: FLT3 and CDK4/6 inhibitors: Signaling mechanisms and tumor burden in subcutaneous and orthotopic mouse models of acute myeloid leukemia

doi: 10.1007/s10928-014-9393-x

Figure Lengend Snippet: Parameter estimates, inter-animal variability (IIV as CV%) and corresponding relative standard errors (%RSE) for the plasma PK-cellular signaling-tumor burden model with pooled data from AMG925 and sorafenib studies

Article Snippet: Lysates were prepared and analyzed for STAT5 Y694 phosphorylation and Rb S780 phosphorylation using MSD (Meso Scale Discovery) assays, which were then normalized to total STAT5 and p38 MAPK values to produce the values of phosphorylated STAT5 (pSTAT5) and of phosphorylated Rb (pRb).

Techniques: Clinical Proteomics, Inhibition, Phospho-proteomics