Structured Review

Cell Signaling Technology Inc phospho egfr tyr1068
Phospho Egfr Tyr1068, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 57 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/phospho egfr tyr1068/product/Cell Signaling Technology Inc
Average 99 stars, based on 57 article reviews
Price from $9.99 to $1999.99
phospho egfr tyr1068 - by Bioz Stars, 2020-11
99/100 stars

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Related Articles

Western Blot:

Article Title: The enhanced tumor inhibitory effects of gefitinib and L-ascorbic acid combination therapy in non-small cell lung cancer cells
Article Snippet: .. For the western blot analysis, antibodies against EGFR (cat. no. 4267S; dilution, 1:1,000), phosphorylated (p)-EGFR [Tyr845 (cat. no. 6963S; dilution, 1:1,000), Tyr992 (cat. no. 2235S; dilution, 1:1,000) and Tyr1068 (cat. no. 2234S; dilution, 1:1,000)], Akt (cat. no. 9272S; dilution, 1:1,000), p-Akt (cat. no. 4060S; dilution, 1:1,000), signal transducer and activator of transcription 3 (Stat3; cat. no. 9139S; dilution, 1:1,000) and p-Stat3 (cat. no. 9145S; dilution, 1:1,000) were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). .. Antibodies against extracellular signal-related kinase (Erk; cat. no. sc-94; dilution, 1:1,000), p-Erk (cat. no. sc-7383; dilution, 1:1,000) and β-actin (cat. no. sc-47778; dilution, 1:2,000) were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA).

Incubation:

Article Title: PLA Electrospun Scaffolds for Three-Dimensional Triple-Negative Breast Cancer Cell Culture
Article Snippet: .. Transferred membranes were blocked (blocking buffer of 5% bovine serum albumin (BSA) in tris-buffered salineTBS 0.05% Tween (TBS-T)) at room temperature for 1 h. Then, membranes were incubated overnight with the following primary antibodies: Rabbit polyclonal antibodies against pEGFRY1068 (Cell Signaling Technology Inc.; #2234S; dilution 1:1000), STAT3 (Cell Signaling Technology Inc.; #4904S; dilution 1:1000), pSTAT3Y705 (Cell Signaling Technology Inc.; #9131S; dilution 1:1000), rabbit monoclonal antibody against EGFR (Cell Signaling Technology Inc.; #4267S; dilution 1:1000), and mouse polyclonal antibody GAPDH (Proteintech, Manchester, UK; #60004-1-IG; dilution 1:50,000). .. Specific horseradish peroxidase (HRP)-conjugated secondary antibody was incubated for 1 h at room temperature.

other:

Article Title: Discovery and characterization of a novel irreversible EGFR mutants selective and potent kinase inhibitor CHMFL-EGFR-26 with a distinct binding mode
Article Snippet: Phospho-EGF Receptor, Phospho-EGF Receptor (Tyr1068), Stat3, Phospho-Stat3 (Tyr705), AKT, Phospho-AKT (Ser473), Phospho-AKT (Thr308), p44/42 MAPK (Erk1/2), Phospho-p44/42 MAPK (Erk1/2) (Thr202/ Tyr204), p70 S6 Kinase, Phospho-p70 S6 Kinase (Thr389), eIF4E, Phospho-eIF4E (Ser209), 4E-BP1, Phospho-4E-BP1 (Thr37/46) antibody (Cell Signaling Technology) were used for immunoblotting.

Article Title: Docosahexaenoic acid inhibits the phosphorylation of STAT3 and the growth and invasion of renal cancer cells
Article Snippet: Antibodies against epidermal growth factor receptor (EGFR; cat. no. 4267S), phosphorylated (p)-EGFR (p-Tyr1068; cat. no. 2234S), STAT3 (cat. no. 9132S), p-STAT3 (p-Tyr705; cat. no. 9145 L), extracellular signal-regulated kinase (ERK; cat. no. 9102), p-ERK (p-Thr202/Tyr204; cat. no. 9101S), Akt (cat. no. 9272) and p-Akt (p-Ser473; cat. no. 9271; all from Cell Signaling Technology, Inc., Danvers, MA, USA) were used in the present study.

Blocking Assay:

Article Title: PLA Electrospun Scaffolds for Three-Dimensional Triple-Negative Breast Cancer Cell Culture
Article Snippet: .. Transferred membranes were blocked (blocking buffer of 5% bovine serum albumin (BSA) in tris-buffered salineTBS 0.05% Tween (TBS-T)) at room temperature for 1 h. Then, membranes were incubated overnight with the following primary antibodies: Rabbit polyclonal antibodies against pEGFRY1068 (Cell Signaling Technology Inc.; #2234S; dilution 1:1000), STAT3 (Cell Signaling Technology Inc.; #4904S; dilution 1:1000), pSTAT3Y705 (Cell Signaling Technology Inc.; #9131S; dilution 1:1000), rabbit monoclonal antibody against EGFR (Cell Signaling Technology Inc.; #4267S; dilution 1:1000), and mouse polyclonal antibody GAPDH (Proteintech, Manchester, UK; #60004-1-IG; dilution 1:50,000). .. Specific horseradish peroxidase (HRP)-conjugated secondary antibody was incubated for 1 h at room temperature.

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  • 85
    Cell Signaling Technology Inc rabbit polyclonal anti phospho egfr tyr1068
    <t>EGFR</t> activation and Erlotinib antitumor activity in LCSC-derived ADC xenografts. ( a ) Immunoblot analysis of EGFR <t>tyr1068</t> in sensitive (LCSC5) or resistant (LCSC7) LCSCs and in the corresponding xenografts untreated (−) or treated (+) with erlotinib. ( b ) Growth curves of the same control or erlotinib-treated xenografts as in ( a ). Mean±S.D. of three independent experiments is shown. *** P
    Rabbit Polyclonal Anti Phospho Egfr Tyr1068, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 85/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti phospho egfr tyr1068/product/Cell Signaling Technology Inc
    Average 85 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti phospho egfr tyr1068 - by Bioz Stars, 2020-11
    85/100 stars
      Buy from Supplier

    99
    Cell Signaling Technology Inc phospho egfr
    Effects of BLM and A5 on epidermal growth factor receptor <t>(EGFR)</t> and phosphorylated-EGFR in A549 and HCT116 cells. Cells were seeded in 10-cm dishes for 24 h followed by treating with serial concentrations of BLM and A5 (0, 10, 20, 40, 80 μM) for another 24 h. The cells were then harvested for immunoblot analysis as described in the Materials and Methods section. The numbers underneath the blots represent band intensity (normalized to <t>β-Actin,</t> the means of three independent experiments) measured by Image J software. The standard deviations (all within ±15% of the means) were not shown. β-Actin was served as an equal loading control. The experiments were repeated three times.
    Phospho Egfr, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 175 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/phospho egfr/product/Cell Signaling Technology Inc
    Average 99 stars, based on 175 article reviews
    Price from $9.99 to $1999.99
    phospho egfr - by Bioz Stars, 2020-11
    99/100 stars
      Buy from Supplier

    Image Search Results


    EGFR activation and Erlotinib antitumor activity in LCSC-derived ADC xenografts. ( a ) Immunoblot analysis of EGFR tyr1068 in sensitive (LCSC5) or resistant (LCSC7) LCSCs and in the corresponding xenografts untreated (−) or treated (+) with erlotinib. ( b ) Growth curves of the same control or erlotinib-treated xenografts as in ( a ). Mean±S.D. of three independent experiments is shown. *** P

    Journal: Cell Death & Disease

    Article Title: Tyr1068-phosphorylated epidermal growth factor receptor (EGFR) predicts cancer stem cell targeting by erlotinib in preclinical models of wild-type EGFR lung cancer

    doi: 10.1038/cddis.2015.217

    Figure Lengend Snippet: EGFR activation and Erlotinib antitumor activity in LCSC-derived ADC xenografts. ( a ) Immunoblot analysis of EGFR tyr1068 in sensitive (LCSC5) or resistant (LCSC7) LCSCs and in the corresponding xenografts untreated (−) or treated (+) with erlotinib. ( b ) Growth curves of the same control or erlotinib-treated xenografts as in ( a ). Mean±S.D. of three independent experiments is shown. *** P

    Article Snippet: Rabbit polyclonal anti-Phospho-EGFR (Tyr1068), -Phospho-EGFR (tyr1173), -Phospho-Akt (Ser473) -Akt, -Caspase-3 (8G10) and -Phospho-Stat3 (Ser727) and mouse monoclonal anti-STAT3 were purchased from Cell Signaling (Beverly, MA, USA).

    Techniques: Activation Assay, Activity Assay, Derivative Assay

    Effects of BLM and A5 on epidermal growth factor receptor (EGFR) and phosphorylated-EGFR in A549 and HCT116 cells. Cells were seeded in 10-cm dishes for 24 h followed by treating with serial concentrations of BLM and A5 (0, 10, 20, 40, 80 μM) for another 24 h. The cells were then harvested for immunoblot analysis as described in the Materials and Methods section. The numbers underneath the blots represent band intensity (normalized to β-Actin, the means of three independent experiments) measured by Image J software. The standard deviations (all within ±15% of the means) were not shown. β-Actin was served as an equal loading control. The experiments were repeated three times.

    Journal: Molecules

    Article Title: Pingyangmycin and Bleomycin Share the Same Cytotoxicity Pathway

    doi: 10.3390/molecules21070862

    Figure Lengend Snippet: Effects of BLM and A5 on epidermal growth factor receptor (EGFR) and phosphorylated-EGFR in A549 and HCT116 cells. Cells were seeded in 10-cm dishes for 24 h followed by treating with serial concentrations of BLM and A5 (0, 10, 20, 40, 80 μM) for another 24 h. The cells were then harvested for immunoblot analysis as described in the Materials and Methods section. The numbers underneath the blots represent band intensity (normalized to β-Actin, the means of three independent experiments) measured by Image J software. The standard deviations (all within ±15% of the means) were not shown. β-Actin was served as an equal loading control. The experiments were repeated three times.

    Article Snippet: Antibodies for cyclin B1, p21, Bax, EGFR, Bcl-2, PARP, β-Actin, cdc25C, Phospho-EGFR (P-EGFR, Tyr1068) was from Cell Signaling Technology (Beverly, MA, USA); PI/RNase was from BD Biosciences (San Diego CA, USA); BCA protein assay kit was from Beyotime Biotechnology Co. Ltd. (Shanghai, China).

    Techniques: Software

    CL4 impairs Matrigel-induced EGFR-integrin αvβ3 interaction. ( a ) BT-549 cells grown in 2D or on Matrigel in the absence or in the presence of 200 nmol/l CL4 or CL4Sc or 10 μmol/l erlotinib for 24 hours were fixed, permeabilized and labelled with anti-αvβ3 LM609 (red) and anti-EGFR (green) antibodies. Co-localization results appear yellow in the merged images. Nuclei were stained with DAPI. All digital images were captured at the same setting to allow direct comparison of staining patterns (Magnification 63×, 0.7× digital zoom). Scale bar = 20 μm. White square indicate the area showed in insets. Arrowheads indicate some co-localization points between EGFR and integrin αvβ3. ( b ) Equal amounts of lysates from BT-549 cells grown in 2D or on Matrigel in the presence of 200 nmol/l CL4 or CL4Sc were directly subjected to Western blotting or prior immunoprecipitated with anti-EGFR antibody. Filter was cut in two pieces that were immunoblotted with anti-EGFR and anti-integrin β3 antibodies, stripped, rejoined and immunoblotted with anti-integrin αv antibody. Dashed line indicates the boundary between the two pieces of the filter. Vinculin was used as a loading control. Molecular weights of indicated proteins are reported ( left ). The bands were quantified by densitometric analysis and the amount of αv or β3 co-immunoprecipitated with EGFR relative to immunoprecipitated EGFR levels is reported. Values are shown relative to CL4Sc control, arbitrarily set to 1 ( right ). The data shown here represent three experiments exhibiting similar effects. Note that no significant change in αv, β3 and EGFR levels was observed in total lysates following CL4 treatment.

    Journal: Scientific Reports

    Article Title: Aptamer-mediated impairment of EGFR-integrin αvβ3 complex inhibits vasculogenic mimicry and growth of triple-negative breast cancers

    doi: 10.1038/srep46659

    Figure Lengend Snippet: CL4 impairs Matrigel-induced EGFR-integrin αvβ3 interaction. ( a ) BT-549 cells grown in 2D or on Matrigel in the absence or in the presence of 200 nmol/l CL4 or CL4Sc or 10 μmol/l erlotinib for 24 hours were fixed, permeabilized and labelled with anti-αvβ3 LM609 (red) and anti-EGFR (green) antibodies. Co-localization results appear yellow in the merged images. Nuclei were stained with DAPI. All digital images were captured at the same setting to allow direct comparison of staining patterns (Magnification 63×, 0.7× digital zoom). Scale bar = 20 μm. White square indicate the area showed in insets. Arrowheads indicate some co-localization points between EGFR and integrin αvβ3. ( b ) Equal amounts of lysates from BT-549 cells grown in 2D or on Matrigel in the presence of 200 nmol/l CL4 or CL4Sc were directly subjected to Western blotting or prior immunoprecipitated with anti-EGFR antibody. Filter was cut in two pieces that were immunoblotted with anti-EGFR and anti-integrin β3 antibodies, stripped, rejoined and immunoblotted with anti-integrin αv antibody. Dashed line indicates the boundary between the two pieces of the filter. Vinculin was used as a loading control. Molecular weights of indicated proteins are reported ( left ). The bands were quantified by densitometric analysis and the amount of αv or β3 co-immunoprecipitated with EGFR relative to immunoprecipitated EGFR levels is reported. Values are shown relative to CL4Sc control, arbitrarily set to 1 ( right ). The data shown here represent three experiments exhibiting similar effects. Note that no significant change in αv, β3 and EGFR levels was observed in total lysates following CL4 treatment.

    Article Snippet: Filters were probed with the indicated primary antibodies: anti-E-cadherin, anti-Vimentin, anti-PDGFRβ, anti-phospho-EGFR (Tyr1068, indicated as pEGFR), anti-EGFR (intracellular), anti-integrin β3 (D7X3 P), anti-integrin αv (D2N5H) (Cell Signaling Technology Inc.); anti-PARP (H-250), anti-α-tubulin (TU-02), anti-vinculin (N-19) (Santa Cruz Biotechnology Inc.); anti-Ku-80 (AHP317; Serotec, Oxford, UK).

    Techniques: Staining, Western Blot, Immunoprecipitation

    The anti-EGFR aptamer blocks the endothelial trans-differentiation of BT-549 cells. ( a ) BT-549 cells, seeded on Matrigel in the presence of 200 nmol/l CL4 or CL4Sc for 24 hours, were stained with anti-VE-cadherin antibody, visualized by fluorescence microscopy and photographed; nuclei were stained with DAPI. Magnification 20×, scale bar = 100 μm. ( b–f ) BT-549 cells were left untreated or treated as in ( a ) and mRNA levels of the indicated VM-genes were determined by RT-qPCR. Bars depict means ± SD of three independent experiments. *** P

    Journal: Scientific Reports

    Article Title: Aptamer-mediated impairment of EGFR-integrin αvβ3 complex inhibits vasculogenic mimicry and growth of triple-negative breast cancers

    doi: 10.1038/srep46659

    Figure Lengend Snippet: The anti-EGFR aptamer blocks the endothelial trans-differentiation of BT-549 cells. ( a ) BT-549 cells, seeded on Matrigel in the presence of 200 nmol/l CL4 or CL4Sc for 24 hours, were stained with anti-VE-cadherin antibody, visualized by fluorescence microscopy and photographed; nuclei were stained with DAPI. Magnification 20×, scale bar = 100 μm. ( b–f ) BT-549 cells were left untreated or treated as in ( a ) and mRNA levels of the indicated VM-genes were determined by RT-qPCR. Bars depict means ± SD of three independent experiments. *** P

    Article Snippet: Filters were probed with the indicated primary antibodies: anti-E-cadherin, anti-Vimentin, anti-PDGFRβ, anti-phospho-EGFR (Tyr1068, indicated as pEGFR), anti-EGFR (intracellular), anti-integrin β3 (D7X3 P), anti-integrin αv (D2N5H) (Cell Signaling Technology Inc.); anti-PARP (H-250), anti-α-tubulin (TU-02), anti-vinculin (N-19) (Santa Cruz Biotechnology Inc.); anti-Ku-80 (AHP317; Serotec, Oxford, UK).

    Techniques: Staining, Fluorescence, Microscopy, Quantitative RT-PCR

    CL4 inhibits αvβ3-dependent cell adhesion to vitronectin. ( a,b ) Cells were mock-treated or pretreated with 200 nmol/l CL4 or CL4Sc, 10 μmol/l erlotinib, 100 μmol/l cetuximab or 10 μg/ml anti-αvβ3 LM609, as indicated, and then subjected to the adhesion assay on vitronectin-coated plates. Results are expressed as percentage of adherent cells considering the mock-treated control cells as 100%. In b (insert), lysates from NIH3T3 cells were immunoblotted with anti-integrin β3, anti-EGFR and α-tubulin antibodies. ( c,d ) Representative phase-contrast images of BT-549 cells grown on Matrigel monolayer for the indicated times in the absence or in the presence of 200 nmol/l CL4 or 100 μmol/l cetuximab ( c ) or 10 μg/ml anti-αvβ3 LM609 ( d ). Magnification 10×, scale bar = 200 μm. Tube formation ability was determined as the percentage of reduction in loop formation of treated cells compared with mock-treated cells. Note that in the presence of CL4 treatment no loops were observed. Bars depict means ± SD of three independent experiments. *** P

    Journal: Scientific Reports

    Article Title: Aptamer-mediated impairment of EGFR-integrin αvβ3 complex inhibits vasculogenic mimicry and growth of triple-negative breast cancers

    doi: 10.1038/srep46659

    Figure Lengend Snippet: CL4 inhibits αvβ3-dependent cell adhesion to vitronectin. ( a,b ) Cells were mock-treated or pretreated with 200 nmol/l CL4 or CL4Sc, 10 μmol/l erlotinib, 100 μmol/l cetuximab or 10 μg/ml anti-αvβ3 LM609, as indicated, and then subjected to the adhesion assay on vitronectin-coated plates. Results are expressed as percentage of adherent cells considering the mock-treated control cells as 100%. In b (insert), lysates from NIH3T3 cells were immunoblotted with anti-integrin β3, anti-EGFR and α-tubulin antibodies. ( c,d ) Representative phase-contrast images of BT-549 cells grown on Matrigel monolayer for the indicated times in the absence or in the presence of 200 nmol/l CL4 or 100 μmol/l cetuximab ( c ) or 10 μg/ml anti-αvβ3 LM609 ( d ). Magnification 10×, scale bar = 200 μm. Tube formation ability was determined as the percentage of reduction in loop formation of treated cells compared with mock-treated cells. Note that in the presence of CL4 treatment no loops were observed. Bars depict means ± SD of three independent experiments. *** P

    Article Snippet: Filters were probed with the indicated primary antibodies: anti-E-cadherin, anti-Vimentin, anti-PDGFRβ, anti-phospho-EGFR (Tyr1068, indicated as pEGFR), anti-EGFR (intracellular), anti-integrin β3 (D7X3 P), anti-integrin αv (D2N5H) (Cell Signaling Technology Inc.); anti-PARP (H-250), anti-α-tubulin (TU-02), anti-vinculin (N-19) (Santa Cruz Biotechnology Inc.); anti-Ku-80 (AHP317; Serotec, Oxford, UK).

    Techniques: Cell Adhesion Assay

    CL4 decreases IntegriSense signal in tumors and inhibits VM. ( a , b ) In vivo imaging and quantification of IntegriSense in tumor-bearing mice. ( a ) Representative volume renderings taken at the same color gating from CL4Sc- and CL4-treated mice injected with IntegriSense at day 21 (upper panels). Representative images of the single tumors excised from CL4Sc- and CL4-treated mice after in vivo imaging (lower panels). ( b ) The amount of fluorescence (pmoles) was quantified in specific ROIs encompassing the tumor in the animal and normalized to tumor volume (cm 3 ). Error bars depict means ± SD. P = 0.0091 (n = 4). ( c ) Representative sections of tumors (CL4Sc group) were stained with anti-integrin β3 (red) and anti-EGFR (green) antibodies and analysed by confocal microscopy. Nuclei were stained with DAPI. Co-localization results appear yellow in the merged image. Magnification 63×, scale bar = 10 μm. ( d–f ) Equal amounts of lysates from recovered tumors were immunoprecipitated with anti-integrin αvβ3 LM609 antibody ( d ) or anti-EGFR antibody ( e ) and immunoblotted with the indicated antibodies. Total lysates were immunoblotted with anti-EGFR, anti-integrin αv and anti-integrin β3 antibodies, as indicated. Equal loading was confirmed by immunoblot with anti-Ku-80 antibody ( f ). Molecular weights of indicated proteins are reported. Representative data are shown from one of three independent experiments. In d , the histogram reports the amount of EGFR co-immunoprecipitated with integrin αvβ3 relative to αv levels. Values are shown relative to CL4Sc control, arbitrarily set to 1. ( g,h ) Representative sections of tumors harvested from CL4Sc and CL4 groups were stained with integrin β3 ( g ) and PAS/CD31 ( h ) as indicated. (Magnification 40×, scale bar = 50 μm; Magnification 20×, scale bar = 100 μm).

    Journal: Scientific Reports

    Article Title: Aptamer-mediated impairment of EGFR-integrin αvβ3 complex inhibits vasculogenic mimicry and growth of triple-negative breast cancers

    doi: 10.1038/srep46659

    Figure Lengend Snippet: CL4 decreases IntegriSense signal in tumors and inhibits VM. ( a , b ) In vivo imaging and quantification of IntegriSense in tumor-bearing mice. ( a ) Representative volume renderings taken at the same color gating from CL4Sc- and CL4-treated mice injected with IntegriSense at day 21 (upper panels). Representative images of the single tumors excised from CL4Sc- and CL4-treated mice after in vivo imaging (lower panels). ( b ) The amount of fluorescence (pmoles) was quantified in specific ROIs encompassing the tumor in the animal and normalized to tumor volume (cm 3 ). Error bars depict means ± SD. P = 0.0091 (n = 4). ( c ) Representative sections of tumors (CL4Sc group) were stained with anti-integrin β3 (red) and anti-EGFR (green) antibodies and analysed by confocal microscopy. Nuclei were stained with DAPI. Co-localization results appear yellow in the merged image. Magnification 63×, scale bar = 10 μm. ( d–f ) Equal amounts of lysates from recovered tumors were immunoprecipitated with anti-integrin αvβ3 LM609 antibody ( d ) or anti-EGFR antibody ( e ) and immunoblotted with the indicated antibodies. Total lysates were immunoblotted with anti-EGFR, anti-integrin αv and anti-integrin β3 antibodies, as indicated. Equal loading was confirmed by immunoblot with anti-Ku-80 antibody ( f ). Molecular weights of indicated proteins are reported. Representative data are shown from one of three independent experiments. In d , the histogram reports the amount of EGFR co-immunoprecipitated with integrin αvβ3 relative to αv levels. Values are shown relative to CL4Sc control, arbitrarily set to 1. ( g,h ) Representative sections of tumors harvested from CL4Sc and CL4 groups were stained with integrin β3 ( g ) and PAS/CD31 ( h ) as indicated. (Magnification 40×, scale bar = 50 μm; Magnification 20×, scale bar = 100 μm).

    Article Snippet: Filters were probed with the indicated primary antibodies: anti-E-cadherin, anti-Vimentin, anti-PDGFRβ, anti-phospho-EGFR (Tyr1068, indicated as pEGFR), anti-EGFR (intracellular), anti-integrin β3 (D7X3 P), anti-integrin αv (D2N5H) (Cell Signaling Technology Inc.); anti-PARP (H-250), anti-α-tubulin (TU-02), anti-vinculin (N-19) (Santa Cruz Biotechnology Inc.); anti-Ku-80 (AHP317; Serotec, Oxford, UK).

    Techniques: In Vivo Imaging, Mouse Assay, Injection, Fluorescence, Staining, Confocal Microscopy, Immunoprecipitation

    CL4 prevents TNBC cells ability to form VM channels on Matrigel and destroys preformed VM. ( a ) MDA-MB-231 and BT-549 cells were serum-starved for 18 hours and then left untreated or stimulated with 20 ng/ml EGF in the absence or in the presence of 10 μmol/l erlotinib or 200 nmol/l CL4 or CL4Sc, for 15 minutes, as indicated. Cell lysates were immunoblotted with anti-pEGFR and anti-EGFR antibodies. Equal loading was confirmed by immunoblot with anti-vinculin antibody. Values below the blot indicate the ratio of pEGFR to total EGFR signal levels, normalized to the respective vinculin signal level, and reported as relative to EGF stimulated cells in the presence of CL4Sc, arbitrarily set to 1 (labeled with asterisk). ( b ) MDA-MB-231 and BT-549 cells were seeded on Matrigel monolayer in the absence or in the presence of 200 nmol/l CL4 or CL4Sc or 10 μmol/l erlotinib for 24 hours. ( c ) BT-549 cells were seeded on Matrigel, in the presence of 200 nmol/l CL4 or CL4Sc aptamers, for 4 hours. Note that just 4 hours CL4-treatment is sufficient to prevent VM channels formation. ( d ) Lysates from MDA-MB-231 and BT-549 cells grown on Matrigel for 24 hours, in the presence of CL4Sc or CL4, as in ( b ), were immunoblotted with anti-PARP antibody. Vinculin was used as a loading control. The 24 hours-treatment of BT-549 cells with 10 nmol/l trabectedin, an antitumor drug, was used as a positive control of PARP cleavage. ( e ) BT-549 cells were grown on Matrigel for 24 hours and then treated with 200 nmol/l CL4 or CL4Sc for 4 hours. Destruction of preformed tubes was determined as the percentage of intact loops of CL4-treated cells compared with control. ( b,c,e ) Cells were photographed by phase-contrast microscopy. Representative photographs of at least five independent experiments were shown. Magnification 10×, scale bar = 200 μm. Bars depict means ± SD. *** P

    Journal: Scientific Reports

    Article Title: Aptamer-mediated impairment of EGFR-integrin αvβ3 complex inhibits vasculogenic mimicry and growth of triple-negative breast cancers

    doi: 10.1038/srep46659

    Figure Lengend Snippet: CL4 prevents TNBC cells ability to form VM channels on Matrigel and destroys preformed VM. ( a ) MDA-MB-231 and BT-549 cells were serum-starved for 18 hours and then left untreated or stimulated with 20 ng/ml EGF in the absence or in the presence of 10 μmol/l erlotinib or 200 nmol/l CL4 or CL4Sc, for 15 minutes, as indicated. Cell lysates were immunoblotted with anti-pEGFR and anti-EGFR antibodies. Equal loading was confirmed by immunoblot with anti-vinculin antibody. Values below the blot indicate the ratio of pEGFR to total EGFR signal levels, normalized to the respective vinculin signal level, and reported as relative to EGF stimulated cells in the presence of CL4Sc, arbitrarily set to 1 (labeled with asterisk). ( b ) MDA-MB-231 and BT-549 cells were seeded on Matrigel monolayer in the absence or in the presence of 200 nmol/l CL4 or CL4Sc or 10 μmol/l erlotinib for 24 hours. ( c ) BT-549 cells were seeded on Matrigel, in the presence of 200 nmol/l CL4 or CL4Sc aptamers, for 4 hours. Note that just 4 hours CL4-treatment is sufficient to prevent VM channels formation. ( d ) Lysates from MDA-MB-231 and BT-549 cells grown on Matrigel for 24 hours, in the presence of CL4Sc or CL4, as in ( b ), were immunoblotted with anti-PARP antibody. Vinculin was used as a loading control. The 24 hours-treatment of BT-549 cells with 10 nmol/l trabectedin, an antitumor drug, was used as a positive control of PARP cleavage. ( e ) BT-549 cells were grown on Matrigel for 24 hours and then treated with 200 nmol/l CL4 or CL4Sc for 4 hours. Destruction of preformed tubes was determined as the percentage of intact loops of CL4-treated cells compared with control. ( b,c,e ) Cells were photographed by phase-contrast microscopy. Representative photographs of at least five independent experiments were shown. Magnification 10×, scale bar = 200 μm. Bars depict means ± SD. *** P

    Article Snippet: Filters were probed with the indicated primary antibodies: anti-E-cadherin, anti-Vimentin, anti-PDGFRβ, anti-phospho-EGFR (Tyr1068, indicated as pEGFR), anti-EGFR (intracellular), anti-integrin β3 (D7X3 P), anti-integrin αv (D2N5H) (Cell Signaling Technology Inc.); anti-PARP (H-250), anti-α-tubulin (TU-02), anti-vinculin (N-19) (Santa Cruz Biotechnology Inc.); anti-Ku-80 (AHP317; Serotec, Oxford, UK).

    Techniques: Multiple Displacement Amplification, Labeling, Positive Control, Microscopy

    Proposed mechanism of action for CL4 aptamer related with integrin αvβ3-EGFR interaction. By binding to EGFR, CL4 aptamer impairs integrin αvβ3-EGFR interaction, causing inhibition of integrin binding to matrix and, in turn, VM.

    Journal: Scientific Reports

    Article Title: Aptamer-mediated impairment of EGFR-integrin αvβ3 complex inhibits vasculogenic mimicry and growth of triple-negative breast cancers

    doi: 10.1038/srep46659

    Figure Lengend Snippet: Proposed mechanism of action for CL4 aptamer related with integrin αvβ3-EGFR interaction. By binding to EGFR, CL4 aptamer impairs integrin αvβ3-EGFR interaction, causing inhibition of integrin binding to matrix and, in turn, VM.

    Article Snippet: Filters were probed with the indicated primary antibodies: anti-E-cadherin, anti-Vimentin, anti-PDGFRβ, anti-phospho-EGFR (Tyr1068, indicated as pEGFR), anti-EGFR (intracellular), anti-integrin β3 (D7X3 P), anti-integrin αv (D2N5H) (Cell Signaling Technology Inc.); anti-PARP (H-250), anti-α-tubulin (TU-02), anti-vinculin (N-19) (Santa Cruz Biotechnology Inc.); anti-Ku-80 (AHP317; Serotec, Oxford, UK).

    Techniques: Binding Assay, Inhibition

    Monitoring MET amplification in circulating tumor DNA during anti-EGFR therapy

    Journal: Cancer discovery

    Article Title: Amplification of the MET receptor drives resistance to anti-EGFR therapies in colorectal cancer

    doi: 10.1158/2159-8290.CD-12-0558

    Figure Lengend Snippet: Monitoring MET amplification in circulating tumor DNA during anti-EGFR therapy

    Article Snippet: Primary antibodies: anti-Actin ( – ), anti-EGR and anti-RAS (F234) were from Santa Cruz Biotechnology, anti-MET (clone 3D4) was from Invitrogen; antibodies against phosphorylated Met (Tyr1234/1235 ), phosphorylated EGFR (Tyr1068 ), phosphorylated ERK (Thr202 /Tyr204 ), phosphorylated AKT (Ser473 ), total AKT, ERK were from Cell Signaling.

    Techniques: Amplification

    MET amplification is selected for in KRAS wild-type CRC samples from patients who developed resistance to anti-EGFR treatment

    Journal: Cancer discovery

    Article Title: Amplification of the MET receptor drives resistance to anti-EGFR therapies in colorectal cancer

    doi: 10.1158/2159-8290.CD-12-0558

    Figure Lengend Snippet: MET amplification is selected for in KRAS wild-type CRC samples from patients who developed resistance to anti-EGFR treatment

    Article Snippet: Primary antibodies: anti-Actin ( – ), anti-EGR and anti-RAS (F234) were from Santa Cruz Biotechnology, anti-MET (clone 3D4) was from Invitrogen; antibodies against phosphorylated Met (Tyr1234/1235 ), phosphorylated EGFR (Tyr1068 ), phosphorylated ERK (Thr202 /Tyr204 ), phosphorylated AKT (Ser473 ), total AKT, ERK were from Cell Signaling.

    Techniques: Amplification

    Whole exome analysis reveals increased MET copy number in CRC samples from patients who developed resistance to anti-EGFR treatment

    Journal: Cancer discovery

    Article Title: Amplification of the MET receptor drives resistance to anti-EGFR therapies in colorectal cancer

    doi: 10.1158/2159-8290.CD-12-0558

    Figure Lengend Snippet: Whole exome analysis reveals increased MET copy number in CRC samples from patients who developed resistance to anti-EGFR treatment

    Article Snippet: Primary antibodies: anti-Actin ( – ), anti-EGR and anti-RAS (F234) were from Santa Cruz Biotechnology, anti-MET (clone 3D4) was from Invitrogen; antibodies against phosphorylated Met (Tyr1234/1235 ), phosphorylated EGFR (Tyr1068 ), phosphorylated ERK (Thr202 /Tyr204 ), phosphorylated AKT (Ser473 ), total AKT, ERK were from Cell Signaling.

    Techniques: