Journal: Carcinogenesis

Article Title: Inhibition of EGFR signaling in human prostate cancer PC-3 cells by combination treatment with beta-phenylethyl isothiocyanate and curcumin.

doi: 10.1093/carcin/bgi272

Figure Lengend Snippet: Fig. 3. Inhibition of EGFR phosphorylation by PEITC, curcumin, and their combination after stimulation of PC-3 C4 cells with EGF. (A) PC-3 C4 cells were stimulated with EGF (100 ng/ml) for 10 min after a 5 min pretreatment with compounds (curcumin, 25 mM; PEITC, 10 mM; or their combination) in serum-free media. Activated EGFR proteins were measured using four different types of phospho-specific antibodies, which recognize specific phosphorylation sites on tyrosine residues, by western blotting. Con, control; C, curcumin; P, PEITC. (B) PC-3 C4 cells were pretreated with compounds (curcumin, 25 mM; PEITC, 10 mM; or their combination) for 5 min and then challenged with EGF (100 ng/ml) for 1 h in serum-free media. Activated Akt proteins were measured by western blotting using phospho-specific antibodies that detect specific phosphorylation sites in Akt. Actin was used to ensure equal protein loading. Con, control; C, curcumin; P, PEITC.

Article Snippet: Rabbit polyclonal antibodies against p-EGFR (Y845, Y992 and Y1068), total EGFR, total poly(ADP-Ribose) polymerase (PARP), cleaved PARP, caspase 3, cleaved caspase 3, p-Akt (S473, S308), total Akt, p-PDK1, p-IkBa, total IkBa and p-phosphatidylinositol 3-kinase (PI3K) (p85) were purchased from Cell Signaling (Beverly, MA).

Techniques: Inhibition, Phospho-proteomics, Western Blot, Control

Journal: Carcinogenesis

Article Title: Inhibition of EGFR signaling in human prostate cancer PC-3 cells by combination treatment with beta-phenylethyl isothiocyanate and curcumin.

doi: 10.1093/carcin/bgi272

Figure Lengend Snippet: Fig. 2. Effects of EGF on EGFR signaling in PC-3 C4 cells. PC-3 C4 cells were incubated with EGF (100 ng/ml) in serum-free media for the indicated times. Representative EGFR signaling proteins (p-EGFR, p-PI3K (p85), p-Akt and p-IkBa) were measured using phospho-specific antibodies, which detect phosphorylation at specific residue. Anti-EGFR, anti-PI3K (p85), anti-Akt and anti-IkBa antibodies were used to determine the total expression levels of each protein. Actin was used to ensure equal protein loading.

Article Snippet: Rabbit polyclonal antibodies against p-EGFR (Y845, Y992 and Y1068), total EGFR, total poly(ADP-Ribose) polymerase (PARP), cleaved PARP, caspase 3, cleaved caspase 3, p-Akt (S473, S308), total Akt, p-PDK1, p-IkBa, total IkBa and p-phosphatidylinositol 3-kinase (PI3K) (p85) were purchased from Cell Signaling (Beverly, MA).

Techniques: Incubation, Phospho-proteomics, Residue, Expressing

Journal: Carcinogenesis

Article Title: Inhibition of EGFR signaling in human prostate cancer PC-3 cells by combination treatment with beta-phenylethyl isothiocyanate and curcumin.

doi: 10.1093/carcin/bgi272

Figure Lengend Snippet: Fig. 4. Additive inhibition of EGFR signaling in PC-3 parental cells by combination of PEITC and curcumin. (A) PC-3 cells were pretreated with compounds (curcumin, 25 mM; PEITC, 10 mM; or their combination) for 5 min and then stimulated with EGF (100 ng/ml). For evaluation of the EGFR signaling proteins, phospho-specific antibodies against the phosphorylated tyrosine residues (Y845, Y992 and Y1068) of EGFR were used. Phospho-p85, a subunit PI3K, was detected using a specific antibody, which can detect p-YXXM motif (Y, tyrosine; X, amino acid; M, methionine). Actin was used as an equal loading control. Relative fold of activation of p-EGFR (Y1068) was measured by densitometry analysis (right). Con, control; C, curcumin; P, PEITC. (B) PC-3 cells were pretreated with compounds for 5 min and then incubated with EGF (100 ng/ml) for 2 h. Phosphorylation of Akt was measured using specific antibody detecting phosphorylation of S473 or T308, by western blotting. Relative fold activation of p-Akt was measured by densitometry (right). Total protein levels of Akt and actin were used for the Akt expression level and protein equal loading controls, respectively. Phosphorylation of IkBa was also measured by western blotting using a phospho-specific antibody. Con, control; C, curcumin; P, PEITC. (C) PC-3 cells were treated with 0.2% DMSO or 10 mM PEITC for 5 min followed by a 10 min incubation with EGF (100 ng/ml). Phosphorylated EGFR (red) and nuclei (blue) were detected by immunofluorescence analysis using anti-p-EGFR (Y1086) and DAPI dye, respectively. Pictures with red and blue color were merged. Magnitude: 400-fold; Con, control.

Article Snippet: Rabbit polyclonal antibodies against p-EGFR (Y845, Y992 and Y1068), total EGFR, total poly(ADP-Ribose) polymerase (PARP), cleaved PARP, caspase 3, cleaved caspase 3, p-Akt (S473, S308), total Akt, p-PDK1, p-IkBa, total IkBa and p-phosphatidylinositol 3-kinase (PI3K) (p85) were purchased from Cell Signaling (Beverly, MA).

Techniques: Inhibition, Control, Activation Assay, Incubation, Phospho-proteomics, Western Blot, Expressing, Immunofluorescence

Journal: Carcinogenesis

Article Title: Inhibition of EGFR signaling in human prostate cancer PC-3 cells by combination treatment with beta-phenylethyl isothiocyanate and curcumin.

doi: 10.1093/carcin/bgi272

Figure Lengend Snippet: Fig. 5. A schematic representation of possible mechanisms for the combined effect of PEITC and curcumin on EGFR signaling and apoptosis in PC-3 cells. Signaling pathways and blocking mechanisms are indicated with green and red colored line, respectively.

Article Snippet: Rabbit polyclonal antibodies against p-EGFR (Y845, Y992 and Y1068), total EGFR, total poly(ADP-Ribose) polymerase (PARP), cleaved PARP, caspase 3, cleaved caspase 3, p-Akt (S473, S308), total Akt, p-PDK1, p-IkBa, total IkBa and p-phosphatidylinositol 3-kinase (PI3K) (p85) were purchased from Cell Signaling (Beverly, MA).

Techniques: Protein-Protein interactions, Blocking Assay

Journal: Therapeutic Advances in Medical Oncology

Article Title: Frequency, impact and a preclinical study of novel ERBB gene family mutations in HER2-positive breast cancer

doi: 10.1177/1758835918778297

Figure Lengend Snippet: The identity and properties of somatic ERBB family mutations confirmed in our Agena study in 227 HER2-positive breast cancers. A low AVSIFT 18 value predicts that a mutation is likely to be deleterious. Mutation Assessor 19 is a bioinformatics tool which predicts the likely effect of a mutation. No mutation reported herein was reported in the TCGA HER2-positive breast cancer study as of 1 April 2017. All ERBB family mutations listed here were confirmed somatic by sequencing matched normal DNA.

Article Snippet: EGFR (Y992) , Cell Signaling , 2235 , Rabbit , 1:100.

Techniques: Mutagenesis, Sequencing

Journal: Therapeutic Advances in Medical Oncology

Article Title: Frequency, impact and a preclinical study of novel ERBB gene family mutations in HER2-positive breast cancer

doi: 10.1177/1758835918778297

Figure Lengend Snippet: Primary antibodies used in our RPPA experiments.

Article Snippet: EGFR (Y992) , Cell Signaling , 2235 , Rabbit , 1:100.

Techniques: