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  • 99
    Millipore gefitinib iressa
    Blockade of EC-pericyte interactions in vivo leads to decreased basement membrane deposition and increased EC vessel width . (A) Quail CAM tissue from the controls, <t>gefitinib/imatinib-treated</t> quail embryos and α-PDGF-BB/HB-EGF–treated embryos, was isolated and immunostained for the basement membrane component fibronectin. Quantification of immunostaining intensity of extracellular basement membrane protein deposition displays a decrease in deposition under conditions of inhibited pericyte recruitment, most severely in conditions of combined PDGFR and EGFR inhibition. (B) Representative images of the fibronectin stains are shown, with arrows highlighting areas of decreased levels of extracellular basement membrane protein deposition. Overlays of QH1 staining (EC marker, green) versus fibronectin (red) are included for control versus α-PDGF-BB/HB-EGF treatments. (C) Measurements of EC tube width were done (from QH1 stains of EC tubes), demonstrating increased EC vessel width under conditions of inhibited pericyte recruitment to EC tubes. Furthermore, there was a decrease in the number of EC branch points in CAMs treated with these chemical inhibitors or blocking antibodies, further implicating direct vascular phenotypes. (D) Representative images of CAM tissue stained with the quail EC specific marker, QH1, are shown demonstrating the increased vessel width and decreased branch point phenotypes. Arrowheads indicate the “membrane-ruffled appearance” that was particularly observed in the <t>gefitinib/imatinib</t> condition and correlated with strongly reduced fibronectin deposition and increased vessel widths. n ≥ 5; P ≤ .01. *Significance from control conditions. +Significance from individual factor addition.
    Gefitinib Iressa, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Tocris gefitinib
    Effects of <t>gefitinib</t> with or without Ad-p53 on the growth of MDA-MB-468 cells. Firstly, cells were infected by Ad-p53 for 24 h; vehicle-treated cells were treated with DMSO. Then, cells with or without Ad-p53 were treated by fixed-ratio concentrations of gefitinib for 48 h, and cell viability was assessed by MTT assay. The results represent means ± SEM from three independent experiments. Ad-p53, recombinant human p53 adenovirus; DMSO, dimethyl sulfoxide.
    Gefitinib, supplied by Tocris, used in various techniques. Bioz Stars score: 95/100, based on 157 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Selleck Chemicals gefitinib
    Prevention of ACC Ar and EGF-induced metastasis in vivo by <t>gefitinib.</t> For in vivo metastasis analyses, the pulmonary metastatic models of mice were established by injected with ACC cells with indicated treatment by tail vein. The ACC-2 cells were pretreated with EGF (10 ng/ml) for 48 h, and gefitinib (1 µM) for 1 h, as well as ACC-2 Ar were used in the assays. ACC-2 parental cells were also injected as Control. A, Representative hematoxylin and eosin-stained histological sections of lungs from the mice injected with ACC cells. B, Quantitative analysis of pulmonary metastasis from mice as described above. Lesions of 0.1 mm or higher were counted from 20 slides per treatment (Mean ± SE, n = 5 for each group, * p
    Gefitinib, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 99/100, based on 654 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    AstraZeneca zd 1839
    Case 4: Brain CT-scan at baseline ( A ) and after 3 months of ZD1839 therapy ( B ). Brain metastasis from NSCLC responding to <t>ZD</t> 1839 therapy. This patient has been pretreated with three lines of chemotherapy including platinum and taxanes, and received ZD 1839 after whole-brain radiotherapy failure.
    Zd 1839, supplied by AstraZeneca, used in various techniques. Bioz Stars score: 88/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    LC Laboratories iressa gefitinib
    SKLB188 inhibits the phosphorylation of EGFR and its downstream MEK/Erk and Akt/mTOR pathways in FaDu cells. ( A and B ) Serum-starved FaDu cells were treated with SKLB188 (0–2 μ M ) or <t>Iressa</t> (0 –2 μ M ) for 24 h, followed by stimulation with EGF (50 ng ml −1 ) for 1 h. The cell lysates were subject to western blotting with indicated antibodies.
    Iressa Gefitinib, supplied by LC Laboratories, used in various techniques. Bioz Stars score: 91/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Sequoia Research gefitinib iressa
    SKLB188 inhibits the phosphorylation of EGFR and its downstream MEK/Erk and Akt/mTOR pathways in FaDu cells. ( A and B ) Serum-starved FaDu cells were treated with SKLB188 (0–2 μ M ) or <t>Iressa</t> (0 –2 μ M ) for 24 h, followed by stimulation with EGF (50 ng ml −1 ) for 1 h. The cell lysates were subject to western blotting with indicated antibodies.
    Gefitinib Iressa, supplied by Sequoia Research, used in various techniques. Bioz Stars score: 91/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    AstraZeneca gefitinib zd1839 iressa
    SKLB188 inhibits the phosphorylation of EGFR and its downstream MEK/Erk and Akt/mTOR pathways in FaDu cells. ( A and B ) Serum-starved FaDu cells were treated with SKLB188 (0–2 μ M ) or <t>Iressa</t> (0 –2 μ M ) for 24 h, followed by stimulation with EGF (50 ng ml −1 ) for 1 h. The cell lysates were subject to western blotting with indicated antibodies.
    Gefitinib Zd1839 Iressa, supplied by AstraZeneca, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Tocris egfr tki gefitinib
    Comparison of Mig-6 expression in lung adenocarcinoma at baseline and after acquiring <t>EGFR-TKI</t> resistance. a–d Representative pictures of Mig-6 by immunohistochemistry in cases 1 and 2. a and c are initial biopsy samples. b and d are matched re-biopsy samples of case 1 and 2. The expression of Mig-6 in re-biopsy samples were significantly higher than those of initial biopsy samples. e and f Analysis of Mig-6 expressions at baseline and after acquiring EGFR-TKI resistance ( n = 26)
    Egfr Tki Gefitinib, supplied by Tocris, used in various techniques. Bioz Stars score: 90/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    AstraZeneca reagents gefitinib iressa
    Effect of combined treatment of erlotinib or <t>gefitinib</t> with MTE on tumor growth, histological changes in HCC827/ER mice xenografts Tumor volume (A) and weight (B) of HCC827/ER xenografts was assessed. Mice were treated for 3 weeks with vehicle (control), gefitinib or erlotinib (50 mg/kg, p.o.), MTE (5 g/kg, i.p.), or combinations of MTE and one of the 2 other drugs as described in Materials and Methods. The weight of resected tumors was measured after animals were sacrificed. Histological changes (C) were detected by HE staining (200 ×) and immunohistochemistry staining to compare tumor growth (PCNA), cell apoptosis (TUNEL), tumor angiogenesis (CD105, VEGF), and EMT makers (E-cadherin, vimentin) between various treatment groups. Data are presented as the mean ± SE from mice in each group (n = 8). *p
    Reagents Gefitinib Iressa, supplied by AstraZeneca, used in various techniques. Bioz Stars score: 91/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Biaffin gefitinib iressa
    Lapatinib attenuates HER2CA GH3 tumor growth and hormone secretion in vivo more than <t>gefitinib.</t> A, GH3 cells stably expressing HER2CA (3 × 10 6 cells per rat, 0.2 ml with matrigel) or pcDNA3 were inoculated sc in WF rats (4–5 wk of age).
    Gefitinib Iressa, supplied by Biaffin, used in various techniques. Bioz Stars score: 89/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Santa Cruz Biotechnology gefitinib iressa
    Sprr2d and Slpi expression in infundibula of K5cre-CMVcaNrf2 mice. A Average number of BrdU-positive cells in infundibula (INF) of tg/wt and tg/tg mice ( N = 4). B,C Average thickness of infundibula and number of PCNA-positive cells in infundibula of P32 tg/wt and tg/tg mice injected with vehicle or <t>Gefitinib</t> ( N = 4/5). D Upper panel: immunohistochemistry of Sprr2 on longitudinal P10 tg/wt and tg/tg back skin sections. Note staining of Sprr2 in differentiated infundibular keratinocytes in tg/tg mice. Scale bar: 25 μm. Middle panel: toluidine blue staining for detection of mast cells on longitudinal back skin sections of P32 tg/wt and tg/tg mice. Note increase in number of mast cells in tg/tg mice and assembly of mast cells along the interfollicular epidermis and upper part of the hair follicles. Scale bar: 100 μm. Lower panel: immunohistochemistry of Slpi on longitudinal P10 tg/wt and tg/tg back skin sections. Inset in lower panel shows immunohistochemistry without primary antibody. The dashed line marks the basement membrane of the interfollicular epidermis. Note staining of Slpi (indicated by arrow) in differentiated infundibular keratinocytes and stratum corneum in tg/tg mice. Scale bar: 25 μm. E Electron microscopy of infundibular stratum corneum of P32 tg/wt and tg/tg mice. Corneocyte layers in lower panel were numbered from basal to distal (C1–C3). Arrows point to corneodesmosomes. Note delayed corneodesmosome degradation in tg/tg mice. C, corneocyte. F Working model: Nrf2 activation leads to upregulation of Slpi, Sprr2d, and Epgn in hair follicle infundibula. Slpi upregulation promotes inhibition of CE protease activity. Consequently, corneodesmosome cleavage is reduced, leading to decreased desquamation and thereby to hyperkeratosis of infundibula. Sprr2d upregulation reduces epidermal barrier functionality, resulting in increased inflammation and consequently stimulation of proliferation. Epgn and other epidermal growth factor (EGF) family members stimulate proliferation of infundibular keratinocytes via EGFR signaling. Together, this results in acanthosis of hair follicle infundibula. Data information: Values are shown as the mean with s.d.
    Gefitinib Iressa, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 89/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Blockade of EC-pericyte interactions in vivo leads to decreased basement membrane deposition and increased EC vessel width . (A) Quail CAM tissue from the controls, gefitinib/imatinib-treated quail embryos and α-PDGF-BB/HB-EGF–treated embryos, was isolated and immunostained for the basement membrane component fibronectin. Quantification of immunostaining intensity of extracellular basement membrane protein deposition displays a decrease in deposition under conditions of inhibited pericyte recruitment, most severely in conditions of combined PDGFR and EGFR inhibition. (B) Representative images of the fibronectin stains are shown, with arrows highlighting areas of decreased levels of extracellular basement membrane protein deposition. Overlays of QH1 staining (EC marker, green) versus fibronectin (red) are included for control versus α-PDGF-BB/HB-EGF treatments. (C) Measurements of EC tube width were done (from QH1 stains of EC tubes), demonstrating increased EC vessel width under conditions of inhibited pericyte recruitment to EC tubes. Furthermore, there was a decrease in the number of EC branch points in CAMs treated with these chemical inhibitors or blocking antibodies, further implicating direct vascular phenotypes. (D) Representative images of CAM tissue stained with the quail EC specific marker, QH1, are shown demonstrating the increased vessel width and decreased branch point phenotypes. Arrowheads indicate the “membrane-ruffled appearance” that was particularly observed in the gefitinib/imatinib condition and correlated with strongly reduced fibronectin deposition and increased vessel widths. n ≥ 5; P ≤ .01. *Significance from control conditions. +Significance from individual factor addition.

    Journal: Blood

    Article Title: Endothelial-derived PDGF-BB and HB-EGF coordinately regulate pericyte recruitment during vasculogenic tube assembly and stabilization

    doi: 10.1182/blood-2010-05-286872

    Figure Lengend Snippet: Blockade of EC-pericyte interactions in vivo leads to decreased basement membrane deposition and increased EC vessel width . (A) Quail CAM tissue from the controls, gefitinib/imatinib-treated quail embryos and α-PDGF-BB/HB-EGF–treated embryos, was isolated and immunostained for the basement membrane component fibronectin. Quantification of immunostaining intensity of extracellular basement membrane protein deposition displays a decrease in deposition under conditions of inhibited pericyte recruitment, most severely in conditions of combined PDGFR and EGFR inhibition. (B) Representative images of the fibronectin stains are shown, with arrows highlighting areas of decreased levels of extracellular basement membrane protein deposition. Overlays of QH1 staining (EC marker, green) versus fibronectin (red) are included for control versus α-PDGF-BB/HB-EGF treatments. (C) Measurements of EC tube width were done (from QH1 stains of EC tubes), demonstrating increased EC vessel width under conditions of inhibited pericyte recruitment to EC tubes. Furthermore, there was a decrease in the number of EC branch points in CAMs treated with these chemical inhibitors or blocking antibodies, further implicating direct vascular phenotypes. (D) Representative images of CAM tissue stained with the quail EC specific marker, QH1, are shown demonstrating the increased vessel width and decreased branch point phenotypes. Arrowheads indicate the “membrane-ruffled appearance” that was particularly observed in the gefitinib/imatinib condition and correlated with strongly reduced fibronectin deposition and increased vessel widths. n ≥ 5; P ≤ .01. *Significance from control conditions. +Significance from individual factor addition.

    Article Snippet: Imatinib was purchased from Cayman Chemical, and gefitinib (Iressa) was purchased from Sigma-Aldrich.

    Techniques: In Vivo, Chick Chorioallantoic Membrane Assay, Isolation, Immunostaining, Inhibition, Staining, Marker, Blocking Assay

    PDGFRβ and EGFR inhibition through the use of chemical inhibitors or neutralizing antibodies in vivo leads to a blockade of pericyte recruitment to EC tubes and concomitant cranial and abdominal hemorrhage phenotypes in developing quail embryos . Two chemical inhibitors and 2 neutralizing antibodies were identified based on their ability to interfere with PDGFR signaling (imatinib and α-PDGF-BB) and EGFR signaling (gefitinib and α-HB-EGF) and administered individually or in combination to quail at 72 hours of embryonic development at a doses of 100nM for the chemical inhibitors and 20 μg/mL for the neutralizing antibodies. The quail were then allowed to develop for 144 hours, at which time the eggs were cracked and the embryos assessed for vascular phenotypes. (A-B) Embryos treated with individual reagents developed mild cranial hemorrhages, while those embryos treated with both gefitinib/imatinib or α-PDGF-BB/HB-EGF, to block PDGFR and EGFR signaling simultaneously, led to more severe hemorrhage phenotypes (Table A). (C-D) CAM tissue from control, gefitinib/imatinib double treatment, and α-PDGF-BB/HB-EGF double treatment embryos was isolated and double stained for the quail EC-specific marker QH1 (green) and PDGFRβ (red). (C) Representative images are shown demonstrating pericyte association with microvascular beds. Arrows denote representative nonassociated pericytes. (D) The number of nonassociated pericytes per high-powered field was quantified, showing an increase in the number of nonassociated pericytes with blood vessels in vivo after treatments to inhibit PDGFR and EGFR signaling. n ≥ 5; P ≤ .01.

    Journal: Blood

    Article Title: Endothelial-derived PDGF-BB and HB-EGF coordinately regulate pericyte recruitment during vasculogenic tube assembly and stabilization

    doi: 10.1182/blood-2010-05-286872

    Figure Lengend Snippet: PDGFRβ and EGFR inhibition through the use of chemical inhibitors or neutralizing antibodies in vivo leads to a blockade of pericyte recruitment to EC tubes and concomitant cranial and abdominal hemorrhage phenotypes in developing quail embryos . Two chemical inhibitors and 2 neutralizing antibodies were identified based on their ability to interfere with PDGFR signaling (imatinib and α-PDGF-BB) and EGFR signaling (gefitinib and α-HB-EGF) and administered individually or in combination to quail at 72 hours of embryonic development at a doses of 100nM for the chemical inhibitors and 20 μg/mL for the neutralizing antibodies. The quail were then allowed to develop for 144 hours, at which time the eggs were cracked and the embryos assessed for vascular phenotypes. (A-B) Embryos treated with individual reagents developed mild cranial hemorrhages, while those embryos treated with both gefitinib/imatinib or α-PDGF-BB/HB-EGF, to block PDGFR and EGFR signaling simultaneously, led to more severe hemorrhage phenotypes (Table A). (C-D) CAM tissue from control, gefitinib/imatinib double treatment, and α-PDGF-BB/HB-EGF double treatment embryos was isolated and double stained for the quail EC-specific marker QH1 (green) and PDGFRβ (red). (C) Representative images are shown demonstrating pericyte association with microvascular beds. Arrows denote representative nonassociated pericytes. (D) The number of nonassociated pericytes per high-powered field was quantified, showing an increase in the number of nonassociated pericytes with blood vessels in vivo after treatments to inhibit PDGFR and EGFR signaling. n ≥ 5; P ≤ .01.

    Article Snippet: Imatinib was purchased from Cayman Chemical, and gefitinib (Iressa) was purchased from Sigma-Aldrich.

    Techniques: Inhibition, In Vivo, Blocking Assay, Chick Chorioallantoic Membrane Assay, Isolation, Staining, Marker

    UA exhibits increased cytotoxicity over conventional chemotherapeutics. ( A, B ) U373MG cells were treated with increasing concentrations of UA (0-200μM), TMZ, BCNU, or Gefitinib (0-500μM) for 48 hours (A) or 6 days (B) and analysed using Alamar blue cell viability assay. Statistical analysis was carried out using non-linear regression analysis and Two-way ANOVA with Bonferroni post-tests, (n=3) (*P

    Journal: bioRxiv

    Article Title: Ursolic acid inhibits cell migration and promotes JNK-dependent lysosomal associated cell death in Glioblastoma multiforme cells

    doi: 10.1101/2020.03.11.987578

    Figure Lengend Snippet: UA exhibits increased cytotoxicity over conventional chemotherapeutics. ( A, B ) U373MG cells were treated with increasing concentrations of UA (0-200μM), TMZ, BCNU, or Gefitinib (0-500μM) for 48 hours (A) or 6 days (B) and analysed using Alamar blue cell viability assay. Statistical analysis was carried out using non-linear regression analysis and Two-way ANOVA with Bonferroni post-tests, (n=3) (*P

    Article Snippet: Cytotoxicity Dose response curves for commonly employed chemotherapeutic drugs used for the treatment of GBM: Temozolomide (TMZ) (Sigma-Aldrich, Arklow, Ireland), Carmustine (BCNU) (Sigma-Aldrich, Arklow, Ireland) and Gefitinib (Insight Biotechnology ltd, Wembley, UK) UA standard (Sigma-Aldrich, Arklow, Ireland) were established.

    Techniques: Viability Assay

    Effects of gefitinib with or without Ad-p53 on the growth of MDA-MB-468 cells. Firstly, cells were infected by Ad-p53 for 24 h; vehicle-treated cells were treated with DMSO. Then, cells with or without Ad-p53 were treated by fixed-ratio concentrations of gefitinib for 48 h, and cell viability was assessed by MTT assay. The results represent means ± SEM from three independent experiments. Ad-p53, recombinant human p53 adenovirus; DMSO, dimethyl sulfoxide.

    Journal: Oncology Reports

    Article Title: Ad-p53 enhances the sensitivity of triple-negative breast cancer MDA-MB-468 cells to the EGFR inhibitor gefitinib

    doi: 10.3892/or.2014.3665

    Figure Lengend Snippet: Effects of gefitinib with or without Ad-p53 on the growth of MDA-MB-468 cells. Firstly, cells were infected by Ad-p53 for 24 h; vehicle-treated cells were treated with DMSO. Then, cells with or without Ad-p53 were treated by fixed-ratio concentrations of gefitinib for 48 h, and cell viability was assessed by MTT assay. The results represent means ± SEM from three independent experiments. Ad-p53, recombinant human p53 adenovirus; DMSO, dimethyl sulfoxide.

    Article Snippet: Gefitinib was obtained from Tocris Bioscience Company (Bristol, UK), minimum purity > 98%, and dissolved in 100% dimethyl sulfoxide (DMSO; Fisher Scientific, Pittsburgh, PA, USA).

    Techniques: Multiple Displacement Amplification, Infection, MTT Assay, Recombinant

    Ad-p53 and gefitinib in combination obviously reduces clonogenic survival. MDA-MB-468 cells were treated with 3 μM of gefitinib alone, MOI of 100 of Ad-p53 alone or in combination for 48 h, and then replaced with new medium. Colonies ( > 50 cells) were counted after being cultured for 14 days. (A) Survival is expressed relative to the untreated controls. The results represent means ± SEM from three independent experiments. Statistical significance was assessed by ANOVA. Tukey’s multiple comparison was applied to compare two subsequent samples. ** P

    Journal: Oncology Reports

    Article Title: Ad-p53 enhances the sensitivity of triple-negative breast cancer MDA-MB-468 cells to the EGFR inhibitor gefitinib

    doi: 10.3892/or.2014.3665

    Figure Lengend Snippet: Ad-p53 and gefitinib in combination obviously reduces clonogenic survival. MDA-MB-468 cells were treated with 3 μM of gefitinib alone, MOI of 100 of Ad-p53 alone or in combination for 48 h, and then replaced with new medium. Colonies ( > 50 cells) were counted after being cultured for 14 days. (A) Survival is expressed relative to the untreated controls. The results represent means ± SEM from three independent experiments. Statistical significance was assessed by ANOVA. Tukey’s multiple comparison was applied to compare two subsequent samples. ** P

    Article Snippet: Gefitinib was obtained from Tocris Bioscience Company (Bristol, UK), minimum purity > 98%, and dissolved in 100% dimethyl sulfoxide (DMSO; Fisher Scientific, Pittsburgh, PA, USA).

    Techniques: Multiple Displacement Amplification, Cell Culture

    Combination of Ad-p53 and gefitinib suppresses the Akt pathway in MDA-MB-468 cells and increases the activity of caspase cascade protein. Cells were treated with Ad-p53 (MOI of 100), gefitinib (3 μM), alone or a combination for 48 h. Cell lysates were analyzed via western blotting using the indicated antibodies. GAPDH was used as a loading control. (A) p53 and EGFR expression was detected in the MDA-MB-468 cells by western blotting. Ad-p53 and gefitinib in combination significantly downregulated p-Akt and upregulated caspase-9 and cleaved caspase-3. ERK and p-ERK showed little change among the four groups. (B) Relative expression of p-Akt, (C) caspase-9 and (D) cleaved caspase-3 was evaluated by ANOVA. Tukey’s multiple comparison was applied to compare two subsequent samples. Data represent means ± SEM from three independent experiments. ** P

    Journal: Oncology Reports

    Article Title: Ad-p53 enhances the sensitivity of triple-negative breast cancer MDA-MB-468 cells to the EGFR inhibitor gefitinib

    doi: 10.3892/or.2014.3665

    Figure Lengend Snippet: Combination of Ad-p53 and gefitinib suppresses the Akt pathway in MDA-MB-468 cells and increases the activity of caspase cascade protein. Cells were treated with Ad-p53 (MOI of 100), gefitinib (3 μM), alone or a combination for 48 h. Cell lysates were analyzed via western blotting using the indicated antibodies. GAPDH was used as a loading control. (A) p53 and EGFR expression was detected in the MDA-MB-468 cells by western blotting. Ad-p53 and gefitinib in combination significantly downregulated p-Akt and upregulated caspase-9 and cleaved caspase-3. ERK and p-ERK showed little change among the four groups. (B) Relative expression of p-Akt, (C) caspase-9 and (D) cleaved caspase-3 was evaluated by ANOVA. Tukey’s multiple comparison was applied to compare two subsequent samples. Data represent means ± SEM from three independent experiments. ** P

    Article Snippet: Gefitinib was obtained from Tocris Bioscience Company (Bristol, UK), minimum purity > 98%, and dissolved in 100% dimethyl sulfoxide (DMSO; Fisher Scientific, Pittsburgh, PA, USA).

    Techniques: Multiple Displacement Amplification, Activity Assay, Western Blot, Expressing

    Ad-p53 and gefitinib combination therapy significantly reduces tumor volume. The analysis of the xenograft tumor volumes was performed according to Materials and methods. (A) Tumor inhibition rate was calculated after MDA-MB-468 cells were treated with Ad-p53 and/or gefitinib. Data represent means ± SEM from three independent experiments. (B) A slight decrease in size was observed when the nude mice were treated with either (b) Ad-p53 or (c) gefitinib. (d) Tumor volume was significantly decreased after Ad-p53 and gefitinib were administered in combination, and (a) the volume of the vehicle-treated xenografts increased slightly. Ad-p53, recombinant human p53 adenovirus.

    Journal: Oncology Reports

    Article Title: Ad-p53 enhances the sensitivity of triple-negative breast cancer MDA-MB-468 cells to the EGFR inhibitor gefitinib

    doi: 10.3892/or.2014.3665

    Figure Lengend Snippet: Ad-p53 and gefitinib combination therapy significantly reduces tumor volume. The analysis of the xenograft tumor volumes was performed according to Materials and methods. (A) Tumor inhibition rate was calculated after MDA-MB-468 cells were treated with Ad-p53 and/or gefitinib. Data represent means ± SEM from three independent experiments. (B) A slight decrease in size was observed when the nude mice were treated with either (b) Ad-p53 or (c) gefitinib. (d) Tumor volume was significantly decreased after Ad-p53 and gefitinib were administered in combination, and (a) the volume of the vehicle-treated xenografts increased slightly. Ad-p53, recombinant human p53 adenovirus.

    Article Snippet: Gefitinib was obtained from Tocris Bioscience Company (Bristol, UK), minimum purity > 98%, and dissolved in 100% dimethyl sulfoxide (DMSO; Fisher Scientific, Pittsburgh, PA, USA).

    Techniques: Inhibition, Multiple Displacement Amplification, Mouse Assay, Recombinant

    Induction of apoptosis following treatments with 3 μM of gefitinib alone, MOI of 100 of Ad-p53 alone or in combination for 48 h. The apoptosis of MDA-MB-468 cells was detected via Annexin V/FITC using flow cytometry. (A) Percentage of apoptotic cells was obtained from UR and LR panels in each scatter plot for all the treatment groups. (B) Data represent means ± SEM from three independent experiments. Statistical significance was assessed by ANOVA. Tukey’s multiple comparison was applied to compare two subsequent samples. ** P

    Journal: Oncology Reports

    Article Title: Ad-p53 enhances the sensitivity of triple-negative breast cancer MDA-MB-468 cells to the EGFR inhibitor gefitinib

    doi: 10.3892/or.2014.3665

    Figure Lengend Snippet: Induction of apoptosis following treatments with 3 μM of gefitinib alone, MOI of 100 of Ad-p53 alone or in combination for 48 h. The apoptosis of MDA-MB-468 cells was detected via Annexin V/FITC using flow cytometry. (A) Percentage of apoptotic cells was obtained from UR and LR panels in each scatter plot for all the treatment groups. (B) Data represent means ± SEM from three independent experiments. Statistical significance was assessed by ANOVA. Tukey’s multiple comparison was applied to compare two subsequent samples. ** P

    Article Snippet: Gefitinib was obtained from Tocris Bioscience Company (Bristol, UK), minimum purity > 98%, and dissolved in 100% dimethyl sulfoxide (DMSO; Fisher Scientific, Pittsburgh, PA, USA).

    Techniques: Multiple Displacement Amplification, Flow Cytometry, Cytometry

    Ad-p53 infection enhances G 2 /M arrest induced by gefitinib. MDA-MB-468 cells were treated with 3 μM of gefitinib alone, MOI of 100 of Ad-p53 alone or in combination. After 48 h, cell cycle distribution was analyzed by flow cytometry at the indicated time. The graph is based on three independent measurements with similar results. Ad-p53, recombinant human p53 adenovirus; MOI, multiplicity of infection.

    Journal: Oncology Reports

    Article Title: Ad-p53 enhances the sensitivity of triple-negative breast cancer MDA-MB-468 cells to the EGFR inhibitor gefitinib

    doi: 10.3892/or.2014.3665

    Figure Lengend Snippet: Ad-p53 infection enhances G 2 /M arrest induced by gefitinib. MDA-MB-468 cells were treated with 3 μM of gefitinib alone, MOI of 100 of Ad-p53 alone or in combination. After 48 h, cell cycle distribution was analyzed by flow cytometry at the indicated time. The graph is based on three independent measurements with similar results. Ad-p53, recombinant human p53 adenovirus; MOI, multiplicity of infection.

    Article Snippet: Gefitinib was obtained from Tocris Bioscience Company (Bristol, UK), minimum purity > 98%, and dissolved in 100% dimethyl sulfoxide (DMSO; Fisher Scientific, Pittsburgh, PA, USA).

    Techniques: Infection, Multiple Displacement Amplification, Flow Cytometry, Cytometry, Recombinant

    Growth control graphs showing effect of selected HER-family TKIs in doubling concentrations on growth of breast cancer cell lines. ( a ) EGFR reversible inhibitor erlotinib. ( b ) Dual EGFR/HER2 reversible inhibitor lapatinib. ( c ) Pan-HER reversible inhibitor sapitinib. ( d ) Pan-HER irreversible inhibitor canertinib. ( e ) Pan-HER irreversible inhibitor afatinib. Sulforhodamine B colorimetric assay was used to determine the effect of treatment of breast cancer cell lines with doubling dilutions of HER-family inhibiting TKIs. The irreversible pan-inhibitors (e.g. afatinib, canertinib, neratinib) were consistently more effective than the reversible dual and pan inhibitors (e.g. lapatinib, sapitinib), which were in turn more effective than the reversible EGFR inhibitors (e.g. erlotinib, gefitinib). Each point is a representative of the mean ± SD of triplicate samples.

    Journal: Scientific Reports

    Article Title: Synergistic effects of various Her inhibitors in combination with IGF-1R, C-MET and Src targeting agents in breast cancer cell lines

    doi: 10.1038/s41598-017-04301-8

    Figure Lengend Snippet: Growth control graphs showing effect of selected HER-family TKIs in doubling concentrations on growth of breast cancer cell lines. ( a ) EGFR reversible inhibitor erlotinib. ( b ) Dual EGFR/HER2 reversible inhibitor lapatinib. ( c ) Pan-HER reversible inhibitor sapitinib. ( d ) Pan-HER irreversible inhibitor canertinib. ( e ) Pan-HER irreversible inhibitor afatinib. Sulforhodamine B colorimetric assay was used to determine the effect of treatment of breast cancer cell lines with doubling dilutions of HER-family inhibiting TKIs. The irreversible pan-inhibitors (e.g. afatinib, canertinib, neratinib) were consistently more effective than the reversible dual and pan inhibitors (e.g. lapatinib, sapitinib), which were in turn more effective than the reversible EGFR inhibitors (e.g. erlotinib, gefitinib). Each point is a representative of the mean ± SD of triplicate samples.

    Article Snippet: Paclitaxel and gemcitabine were acquired from Sigma-Aldrich (Dorset, UK) and Healthcare at Home (UK), respectively, while Iressa (gefitinib) and crizotinib were purchased from Tocris (Avonmouth, UK), respectively.

    Techniques: Colorimetric Assay

    Effects of the double combination of gefitinib or osimertinib with TPX-0005 in EGFR -mutation-positive NSCLC cells. A. Extracts from the PC9 cell line treated with gefitinib (0.05 μM), TPX-0005 (1 μM), or the double combination for 24 h, were analyzed using the indicated antibodies. Similar results were obtained in three independent experiments. B. Representative immunofluorescence images showing total STAT3 and phospho-STAT3 expression and localization (green) in control, gefitinib, TPX-0005 and gefitinib plus TPX-0005 treated PC9 cells. Cell nuclei were stained with DAPI (blue). Right: expression levels and nuclear translocation of phospho-STAT3 were quantified using an ImageStreamX imaging flow cytometer. C. Extracts from PC9, 11–18 and H1975 cell lines treated with osimertinib (0.05, 0.8, and 0.02 μM respectively), TPX-0005 (1 μM), or osimertinib combined with TPX-0005 for 24 h, were analyzed by Western blotting using the indicated antibodies. β-Actin was used as a loading control. Similar results were obtained in three independent experiments. D. Heatmap depicts the mRNA expression of genes (columns) in control cells and after indicated treatments (rows) compared to the average mRNA expression of each gene in all cell lines. Gefitinib was used at 0.05 μM for PC9, osimertinib was used at 0.02 μM for H1975 and TPX-0005 was used at 1 μM for both cell lines. Quantitative RT-PCR time course experiments from two hours to seven days identified five days as the optimal time-point to compare pathway signaling after treatments (data not shown) and subsequent mRNA expression experiments are presented at this time point for consistency. Data were generated from a minimum of three replicates. β-Actin was used to normalize gene expression. E. Extracts from the PC9 and H1975 cell lines transfected with control siRNA or siRNA against STAT3, Src, YAP1, YES and LYN, were analyzed using the indicated antibodies. Similar results were obtained in three independent experiments. F. Extracts from the PC9 and H1975 cell lines treated with dasatinib (50 and 100 μM respectively) or TPX-0005 (1 μM) for 24 h, were analyzed by Western blotting using the indicated antibodies. Similar results were obtained in three independent experiments.

    Journal: EBioMedicine

    Article Title: Common Co-activation of AXL and CDCP1 in EGFR-mutation-positive Non-Small Cell Lung Cancer Associated With Poor Prognosis

    doi: 10.1016/j.ebiom.2018.02.001

    Figure Lengend Snippet: Effects of the double combination of gefitinib or osimertinib with TPX-0005 in EGFR -mutation-positive NSCLC cells. A. Extracts from the PC9 cell line treated with gefitinib (0.05 μM), TPX-0005 (1 μM), or the double combination for 24 h, were analyzed using the indicated antibodies. Similar results were obtained in three independent experiments. B. Representative immunofluorescence images showing total STAT3 and phospho-STAT3 expression and localization (green) in control, gefitinib, TPX-0005 and gefitinib plus TPX-0005 treated PC9 cells. Cell nuclei were stained with DAPI (blue). Right: expression levels and nuclear translocation of phospho-STAT3 were quantified using an ImageStreamX imaging flow cytometer. C. Extracts from PC9, 11–18 and H1975 cell lines treated with osimertinib (0.05, 0.8, and 0.02 μM respectively), TPX-0005 (1 μM), or osimertinib combined with TPX-0005 for 24 h, were analyzed by Western blotting using the indicated antibodies. β-Actin was used as a loading control. Similar results were obtained in three independent experiments. D. Heatmap depicts the mRNA expression of genes (columns) in control cells and after indicated treatments (rows) compared to the average mRNA expression of each gene in all cell lines. Gefitinib was used at 0.05 μM for PC9, osimertinib was used at 0.02 μM for H1975 and TPX-0005 was used at 1 μM for both cell lines. Quantitative RT-PCR time course experiments from two hours to seven days identified five days as the optimal time-point to compare pathway signaling after treatments (data not shown) and subsequent mRNA expression experiments are presented at this time point for consistency. Data were generated from a minimum of three replicates. β-Actin was used to normalize gene expression. E. Extracts from the PC9 and H1975 cell lines transfected with control siRNA or siRNA against STAT3, Src, YAP1, YES and LYN, were analyzed using the indicated antibodies. Similar results were obtained in three independent experiments. F. Extracts from the PC9 and H1975 cell lines treated with dasatinib (50 and 100 μM respectively) or TPX-0005 (1 μM) for 24 h, were analyzed by Western blotting using the indicated antibodies. Similar results were obtained in three independent experiments.

    Article Snippet: 2.2 Chemicals and Reagents Gefitinib was purchased from Tocris Bioscience Company (Bristol, UK).

    Techniques: Mutagenesis, Immunofluorescence, Expressing, Staining, Translocation Assay, Imaging, Flow Cytometry, Cytometry, Western Blot, Quantitative RT-PCR, Generated, Transfection

    Quantitative RT-PCR analysis and phospho- RTK and non-RTK arrays in EGFR -mutation-positive NSCLC cell lines. Effects of genetic and pharmacologic inhibition of STAT3, YAP1 and SFKs in PC9 and H1975 cells treated with an EGFR TKI A. Expression of various genes in EGFR -mutation-positive NSCLC cell lines. Heatmap depicts gene mRNA expression (columns) in different EGFR - mutation-positive NSCLC cell lines (rows) compared to the average mRNA expression of each gene in all cell lines. B. Phospho-RTK profile in PC9 and H1975 cells obtained by human phospho-kinase array. Each membrane contains kinase specific and positive control antibodies spotted in duplicate. Template tables show the location of tyrosine kinase antibody spotted onto human phospho-RTK array. C. Phospho-kinase array profile in PC9 and H1975 cells obtained by human phospho-kinase array. Each membrane contains kinase specific and positive control antibodies spotted in duplicate. Template tables show the location of tyrosine kinase antibody spotted onto human phospho-kinase array. D. PC9 cells were transiently transfected with STAT3, Src, YAP1, YES, LYN or control siRNA (15 pmol/well). 24 h later cells were treated with serial dilutions of gefitinib. Cell viability was assessed by MTT assay after 72 h of treatment. Plots shown are representative of three independent experiments. E. Effects of negative control siRNA or STAT3 , YAP1 , SRC , YES and LYN siRNA on STAT3 , YAP1 , SRC , YES and LYN mRNA expression after 24 h of transfection. The control condition is set at 1 (arbitrary units). F. PC9 and H1975 cells were treated with serial dilutions of gefitinib or osimertinib, and TPX-0005 alone and with their double combinations for 72 h. The cell viability was measured by MTT and the synergy between the drugs was determined using the Chou and Talalay method (Chou and Talalay plot or Fraction affected [Fa] plot). The dotted horizontal line at 1 indicates the line of additive effect. Fa indicates the fractional inhibition for each CoI. The results represent the means of at least three independent experiments. Data are presented as the means ± standard deviation. G. PC9 and H1975 cells grown in six-well plates (1000 cells/well) for 24 h and then left untreated or treated with gefitinib (0.05 μM), osimertinib (0.02 μM), TPX-0005 (1 μM) alone and with their double combinations. After 72 h, media was replaced with fresh media without drugs. After seven more days cells were washed and stained with crystal violet and then photographed. The crystal violet was extracted and assayed by spectrophotometry. Data are means ± standard deviation of three independent experiments. One-way ANOVA test, * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001.

    Journal: EBioMedicine

    Article Title: Common Co-activation of AXL and CDCP1 in EGFR-mutation-positive Non-Small Cell Lung Cancer Associated With Poor Prognosis

    doi: 10.1016/j.ebiom.2018.02.001

    Figure Lengend Snippet: Quantitative RT-PCR analysis and phospho- RTK and non-RTK arrays in EGFR -mutation-positive NSCLC cell lines. Effects of genetic and pharmacologic inhibition of STAT3, YAP1 and SFKs in PC9 and H1975 cells treated with an EGFR TKI A. Expression of various genes in EGFR -mutation-positive NSCLC cell lines. Heatmap depicts gene mRNA expression (columns) in different EGFR - mutation-positive NSCLC cell lines (rows) compared to the average mRNA expression of each gene in all cell lines. B. Phospho-RTK profile in PC9 and H1975 cells obtained by human phospho-kinase array. Each membrane contains kinase specific and positive control antibodies spotted in duplicate. Template tables show the location of tyrosine kinase antibody spotted onto human phospho-RTK array. C. Phospho-kinase array profile in PC9 and H1975 cells obtained by human phospho-kinase array. Each membrane contains kinase specific and positive control antibodies spotted in duplicate. Template tables show the location of tyrosine kinase antibody spotted onto human phospho-kinase array. D. PC9 cells were transiently transfected with STAT3, Src, YAP1, YES, LYN or control siRNA (15 pmol/well). 24 h later cells were treated with serial dilutions of gefitinib. Cell viability was assessed by MTT assay after 72 h of treatment. Plots shown are representative of three independent experiments. E. Effects of negative control siRNA or STAT3 , YAP1 , SRC , YES and LYN siRNA on STAT3 , YAP1 , SRC , YES and LYN mRNA expression after 24 h of transfection. The control condition is set at 1 (arbitrary units). F. PC9 and H1975 cells were treated with serial dilutions of gefitinib or osimertinib, and TPX-0005 alone and with their double combinations for 72 h. The cell viability was measured by MTT and the synergy between the drugs was determined using the Chou and Talalay method (Chou and Talalay plot or Fraction affected [Fa] plot). The dotted horizontal line at 1 indicates the line of additive effect. Fa indicates the fractional inhibition for each CoI. The results represent the means of at least three independent experiments. Data are presented as the means ± standard deviation. G. PC9 and H1975 cells grown in six-well plates (1000 cells/well) for 24 h and then left untreated or treated with gefitinib (0.05 μM), osimertinib (0.02 μM), TPX-0005 (1 μM) alone and with their double combinations. After 72 h, media was replaced with fresh media without drugs. After seven more days cells were washed and stained with crystal violet and then photographed. The crystal violet was extracted and assayed by spectrophotometry. Data are means ± standard deviation of three independent experiments. One-way ANOVA test, * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001.

    Article Snippet: 2.2 Chemicals and Reagents Gefitinib was purchased from Tocris Bioscience Company (Bristol, UK).

    Techniques: Quantitative RT-PCR, Mutagenesis, Inhibition, Expressing, Positive Control, Transfection, MTT Assay, Negative Control, Standard Deviation, Staining, Spectrophotometry

    HER3 siRNA potentiates the effect of gefitinib ( A ) Comparative growth inhibition rates of MKN45, MKN45-HER3-cs and MKN45-HER3.3 cells exposed to gefitinib. Control siRNA- and HER3.3-transfected MKN45 cells (2 × 10 3 cells/well) were grown in DMEM supplemented with 10% FBS in 96-well plates for 48 h and treated with various concentrations of gefitinib for 48 h. HER3.3 potentiates the effect of gefitinib on MKN45 cell growth, as determined by an MTT assay. The results represent the means ± SDs of three independent experiments. ( B ) Structure of gefitinib. Gefitinib treatment of MKN45 cells significantly induces HER3 ( C ) and HER2 ( D ) mRNA expression, and this induction is prevented by HER3.3 (C). Induction of HER2 expression by gefitinib is not prevented by HER3.3 (D). * P

    Journal: Oncotarget

    Article Title: siRNA-mediated inactivation of HER3 improves the antitumour activity and sensitivity of gefitinib in gastric cancer cells

    doi: 10.18632/oncotarget.17526

    Figure Lengend Snippet: HER3 siRNA potentiates the effect of gefitinib ( A ) Comparative growth inhibition rates of MKN45, MKN45-HER3-cs and MKN45-HER3.3 cells exposed to gefitinib. Control siRNA- and HER3.3-transfected MKN45 cells (2 × 10 3 cells/well) were grown in DMEM supplemented with 10% FBS in 96-well plates for 48 h and treated with various concentrations of gefitinib for 48 h. HER3.3 potentiates the effect of gefitinib on MKN45 cell growth, as determined by an MTT assay. The results represent the means ± SDs of three independent experiments. ( B ) Structure of gefitinib. Gefitinib treatment of MKN45 cells significantly induces HER3 ( C ) and HER2 ( D ) mRNA expression, and this induction is prevented by HER3.3 (C). Induction of HER2 expression by gefitinib is not prevented by HER3.3 (D). * P

    Article Snippet: Reagents and antibodies Gefitinib (Iressa, TOCRIS, UK) was dissolved in dimethyl sulfoxide (DMSO, Sigma) prior to the experiments.

    Techniques: Inhibition, Transfection, MTT Assay, Expressing

    Combination of gefitinib and HER3 siRNA inhibits the PI3K/AKT and ERK signalling pathways MKN45 and MKN45-HER3.3 cells were treated with 20 μM gefitinib. After 48 h of treatment, AKT and ERK activation is simultaneously ablated by the combination compared with the single treatments and the control. HER3.3 yields a greater inhibition of p-AKT than gefitinib.

    Journal: Oncotarget

    Article Title: siRNA-mediated inactivation of HER3 improves the antitumour activity and sensitivity of gefitinib in gastric cancer cells

    doi: 10.18632/oncotarget.17526

    Figure Lengend Snippet: Combination of gefitinib and HER3 siRNA inhibits the PI3K/AKT and ERK signalling pathways MKN45 and MKN45-HER3.3 cells were treated with 20 μM gefitinib. After 48 h of treatment, AKT and ERK activation is simultaneously ablated by the combination compared with the single treatments and the control. HER3.3 yields a greater inhibition of p-AKT than gefitinib.

    Article Snippet: Reagents and antibodies Gefitinib (Iressa, TOCRIS, UK) was dissolved in dimethyl sulfoxide (DMSO, Sigma) prior to the experiments.

    Techniques: Activation Assay, Inhibition

    Metformin inhibits the EMT phenotype in vivo (A) Metformin reversed EMT in fibrotic lung tissue, as shown by immunefluorescence staining. Each image depicts a representative immunostaining of a paraffin-embedded section (4 μm) for E-cadherin in green, Vimentin in red and counter-staining with 4′, 6-diamidino-2-phenylindole (DAPI) in blue. Scale bars = 100 μm. (B) Immunohistochemistry was used to examine the expression of E-cadherin and Vimentin in lung tissues from different groups of mice treated as indicated. The area indicated by the square is shown at higher magnification. Scale bars = 150 μm. BLM, bleomycin; Gef, gefitinib; Met, metformin.

    Journal: Oncotarget

    Article Title: Metformin attenuates gefitinib-induced exacerbation of pulmonary fibrosis by inhibition of TGF-β signaling pathway

    doi:

    Figure Lengend Snippet: Metformin inhibits the EMT phenotype in vivo (A) Metformin reversed EMT in fibrotic lung tissue, as shown by immunefluorescence staining. Each image depicts a representative immunostaining of a paraffin-embedded section (4 μm) for E-cadherin in green, Vimentin in red and counter-staining with 4′, 6-diamidino-2-phenylindole (DAPI) in blue. Scale bars = 100 μm. (B) Immunohistochemistry was used to examine the expression of E-cadherin and Vimentin in lung tissues from different groups of mice treated as indicated. The area indicated by the square is shown at higher magnification. Scale bars = 150 μm. BLM, bleomycin; Gef, gefitinib; Met, metformin.

    Article Snippet: Cell-lines and reagents Gefitinib (Iressa) was purchased from Tocris Bioscience and prepared in dimethyl sulfoxide (DMSO) to obtain a stock solution of 10 mM.

    Techniques: In Vivo, Staining, Immunostaining, Immunohistochemistry, Expressing, Mouse Assay

    Metformin inhibits TGF-β signaling pathway in vivo (A) Metformin decreased expression of α-actin in fibrotic lung tissues from different groups treated as indicated. Paraffin-embedded sections (4 μm) from lung tissues were stained for α-actin using immunohistochemistry. The area indicated by the square is shown at higher magnification. Scale bars = 150 μm. (B) Western blotting analyzed the expression of indicated markers on protein extracts obtained from lung tissues, and β-actin was used as a loading control. BLM, bleomycin; Gef, gefitinib; Met, metformin.

    Journal: Oncotarget

    Article Title: Metformin attenuates gefitinib-induced exacerbation of pulmonary fibrosis by inhibition of TGF-β signaling pathway

    doi:

    Figure Lengend Snippet: Metformin inhibits TGF-β signaling pathway in vivo (A) Metformin decreased expression of α-actin in fibrotic lung tissues from different groups treated as indicated. Paraffin-embedded sections (4 μm) from lung tissues were stained for α-actin using immunohistochemistry. The area indicated by the square is shown at higher magnification. Scale bars = 150 μm. (B) Western blotting analyzed the expression of indicated markers on protein extracts obtained from lung tissues, and β-actin was used as a loading control. BLM, bleomycin; Gef, gefitinib; Met, metformin.

    Article Snippet: Cell-lines and reagents Gefitinib (Iressa) was purchased from Tocris Bioscience and prepared in dimethyl sulfoxide (DMSO) to obtain a stock solution of 10 mM.

    Techniques: In Vivo, Expressing, Staining, Immunohistochemistry, Western Blot

    Metformin attenuates fibrosis induced by conditioned medium from TKI-treated lung cancer PC-9 cells or conditioned medium from TKI-resistant PC-9GR cells (A) Immunofluorescence staining showed that metformin decreased expression of α-actin in HFL-1 cells either induced by conditioned medium from TKI-treated lung cancer PC-9 cells or conditioned medium from TKI-resistant PC-9GR cells. The nucleus were stained with 4′, 6-diamidino-2-phenylindole in the merged images. Scale bars: 150 μm. (B) Metformin decreased TKI-induced expression of COL1A1 and α-actin, and inhibited TKI-enhanced expression of pSMAD2, pSMAD3, pSTAT3, pAKT, and dpERK1/2, as shown by western blot assay. Whole cell protein lysates from HFL-1 cells with different treatments were immunoblotted with antibodies as indicated, and β-actin was used to confirm equal gel loading. Similar results were obtained in three independent experiments. P9, PC-9 cells; R9, PC-9GR cells; CM, conditioned medium; Gef, gefitinib; Met, metformin.

    Journal: Oncotarget

    Article Title: Metformin attenuates gefitinib-induced exacerbation of pulmonary fibrosis by inhibition of TGF-β signaling pathway

    doi:

    Figure Lengend Snippet: Metformin attenuates fibrosis induced by conditioned medium from TKI-treated lung cancer PC-9 cells or conditioned medium from TKI-resistant PC-9GR cells (A) Immunofluorescence staining showed that metformin decreased expression of α-actin in HFL-1 cells either induced by conditioned medium from TKI-treated lung cancer PC-9 cells or conditioned medium from TKI-resistant PC-9GR cells. The nucleus were stained with 4′, 6-diamidino-2-phenylindole in the merged images. Scale bars: 150 μm. (B) Metformin decreased TKI-induced expression of COL1A1 and α-actin, and inhibited TKI-enhanced expression of pSMAD2, pSMAD3, pSTAT3, pAKT, and dpERK1/2, as shown by western blot assay. Whole cell protein lysates from HFL-1 cells with different treatments were immunoblotted with antibodies as indicated, and β-actin was used to confirm equal gel loading. Similar results were obtained in three independent experiments. P9, PC-9 cells; R9, PC-9GR cells; CM, conditioned medium; Gef, gefitinib; Met, metformin.

    Article Snippet: Cell-lines and reagents Gefitinib (Iressa) was purchased from Tocris Bioscience and prepared in dimethyl sulfoxide (DMSO) to obtain a stock solution of 10 mM.

    Techniques: Immunofluorescence, Staining, Expressing, Western Blot

    In combination assays, NSCLC cell lines were seeded in 96-well plates at a density of 10 5 /well with culture medium for 24 h. The cell lines were treated with FU (10 μM) alone or FU plus gefitinib (1 μM)/mithramycin A (50 nM) for another 24 h at 37 °C in a 5 % CO 2 atmosphere. After that, WST-1 was added to each well and incubated for 2 h at 37 °C before measuring absorbance at 490 nm with a Multiskan JX Spectrum instrument. The experiments were performed in triplicate. * p

    Journal: BMC Cancer

    Article Title: Epidermal growth factor signals regulate dihydropyrimidine dehydrogenase expression in EGFR-mutated non-small-cell lung cancer

    doi: 10.1186/s12885-016-2392-0

    Figure Lengend Snippet: In combination assays, NSCLC cell lines were seeded in 96-well plates at a density of 10 5 /well with culture medium for 24 h. The cell lines were treated with FU (10 μM) alone or FU plus gefitinib (1 μM)/mithramycin A (50 nM) for another 24 h at 37 °C in a 5 % CO 2 atmosphere. After that, WST-1 was added to each well and incubated for 2 h at 37 °C before measuring absorbance at 490 nm with a Multiskan JX Spectrum instrument. The experiments were performed in triplicate. * p

    Article Snippet: Drugs Gefitinib was purchased from Tocris Bioscience (Bristol, UK).

    Techniques: Incubation

    Apoptosis was evaluated by ELISA using a Cell Death Detection ELISA Plus Kit (Roche Applied Science). To evaluate the effect of gefitinib and mithramycin A on cell apoptosis, we treated cells with gefitinib (1 μM) and mithramycin A (50 nM) for 24 h. Extracted cell lysates were evaluated by ELISA. In wild-type cells (H1299, H1437) there was little effect of the inhibitors on apoptosis. By contrast, apoptosis increased in EGFR-mutated cell lines treated with gefitinib, except for H1975 with the T790M mutation. Mithramycin A increased apoptosis in all EGFR-mutated cell lines. Gef, gefitinib; Mit A, mithramycin A. The experiment was performed in triplicate. *, p

    Journal: BMC Cancer

    Article Title: Epidermal growth factor signals regulate dihydropyrimidine dehydrogenase expression in EGFR-mutated non-small-cell lung cancer

    doi: 10.1186/s12885-016-2392-0

    Figure Lengend Snippet: Apoptosis was evaluated by ELISA using a Cell Death Detection ELISA Plus Kit (Roche Applied Science). To evaluate the effect of gefitinib and mithramycin A on cell apoptosis, we treated cells with gefitinib (1 μM) and mithramycin A (50 nM) for 24 h. Extracted cell lysates were evaluated by ELISA. In wild-type cells (H1299, H1437) there was little effect of the inhibitors on apoptosis. By contrast, apoptosis increased in EGFR-mutated cell lines treated with gefitinib, except for H1975 with the T790M mutation. Mithramycin A increased apoptosis in all EGFR-mutated cell lines. Gef, gefitinib; Mit A, mithramycin A. The experiment was performed in triplicate. *, p

    Article Snippet: Drugs Gefitinib was purchased from Tocris Bioscience (Bristol, UK).

    Techniques: Enzyme-linked Immunosorbent Assay, Mutagenesis

    a PC9 cells were incubated with various concentrations of EGF, and whole cell lysates were extracted and were examined by immunoblot analysis. b, c . Effects of gefitinib and mithramycin A on ERK, Sp1, and DPD in immunoblot analysis. PC9 cells were pretreated with gefitinib (1 μM) or mithramycin A (50 nM) before administration of 10 ng/ml EGF. After treatment, whole cell lysates were extracted and examined by western blotting. Results are representative of two independent experiments. d PC9 cells were stimulated with inhibitors, and nuclear extracts were prepared to detect the Sp1/DNA interaction by a Trans AM Sp1 kit. The experiment was performed in triplicate. *, p -value

    Journal: BMC Cancer

    Article Title: Epidermal growth factor signals regulate dihydropyrimidine dehydrogenase expression in EGFR-mutated non-small-cell lung cancer

    doi: 10.1186/s12885-016-2392-0

    Figure Lengend Snippet: a PC9 cells were incubated with various concentrations of EGF, and whole cell lysates were extracted and were examined by immunoblot analysis. b, c . Effects of gefitinib and mithramycin A on ERK, Sp1, and DPD in immunoblot analysis. PC9 cells were pretreated with gefitinib (1 μM) or mithramycin A (50 nM) before administration of 10 ng/ml EGF. After treatment, whole cell lysates were extracted and examined by western blotting. Results are representative of two independent experiments. d PC9 cells were stimulated with inhibitors, and nuclear extracts were prepared to detect the Sp1/DNA interaction by a Trans AM Sp1 kit. The experiment was performed in triplicate. *, p -value

    Article Snippet: Drugs Gefitinib was purchased from Tocris Bioscience (Bristol, UK).

    Techniques: Incubation, Western Blot

    Prevention of ACC Ar and EGF-induced metastasis in vivo by gefitinib. For in vivo metastasis analyses, the pulmonary metastatic models of mice were established by injected with ACC cells with indicated treatment by tail vein. The ACC-2 cells were pretreated with EGF (10 ng/ml) for 48 h, and gefitinib (1 µM) for 1 h, as well as ACC-2 Ar were used in the assays. ACC-2 parental cells were also injected as Control. A, Representative hematoxylin and eosin-stained histological sections of lungs from the mice injected with ACC cells. B, Quantitative analysis of pulmonary metastasis from mice as described above. Lesions of 0.1 mm or higher were counted from 20 slides per treatment (Mean ± SE, n = 5 for each group, * p

    Journal: PLoS ONE

    Article Title: Epithelial Mesenchymal Transition Is Required for Acquisition of Anoikis Resistance and Metastatic Potential in Adenoid Cystic Carcinoma

    doi: 10.1371/journal.pone.0051549

    Figure Lengend Snippet: Prevention of ACC Ar and EGF-induced metastasis in vivo by gefitinib. For in vivo metastasis analyses, the pulmonary metastatic models of mice were established by injected with ACC cells with indicated treatment by tail vein. The ACC-2 cells were pretreated with EGF (10 ng/ml) for 48 h, and gefitinib (1 µM) for 1 h, as well as ACC-2 Ar were used in the assays. ACC-2 parental cells were also injected as Control. A, Representative hematoxylin and eosin-stained histological sections of lungs from the mice injected with ACC cells. B, Quantitative analysis of pulmonary metastasis from mice as described above. Lesions of 0.1 mm or higher were counted from 20 slides per treatment (Mean ± SE, n = 5 for each group, * p

    Article Snippet: Gefitinib was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques: In Vivo, Mouse Assay, Injection, Staining

    Case 4: Brain CT-scan at baseline ( A ) and after 3 months of ZD1839 therapy ( B ). Brain metastasis from NSCLC responding to ZD 1839 therapy. This patient has been pretreated with three lines of chemotherapy including platinum and taxanes, and received ZD 1839 after whole-brain radiotherapy failure.

    Journal: British Journal of Cancer

    Article Title: ZD 1839 in patients with brain metastases from non-small-cell lung cancer (NSCLC): report of four cases

    doi: 10.1038/sj.bjc.6601116

    Figure Lengend Snippet: Case 4: Brain CT-scan at baseline ( A ) and after 3 months of ZD1839 therapy ( B ). Brain metastasis from NSCLC responding to ZD 1839 therapy. This patient has been pretreated with three lines of chemotherapy including platinum and taxanes, and received ZD 1839 after whole-brain radiotherapy failure.

    Article Snippet: In pretreated non-small-cell lung cancer (NSCLC) patients, phase I and II studies demonstrated that ZD 1839 is active and well tolerated, with a response rate of about 10–15% ( , ; ).

    Techniques: Computed Tomography

    Treatment schema. On cycle 1, calcitriol ( solid vertical black arrow ) was given on days 1 and 15 and weekly thereafter. Dexamethasone ( broken vertical black arrows ) was given orally 12 h prior, at the time of, and 12 h after to each dose calcitriol. Gefitinib

    Journal: Cancer chemotherapy and pharmacology

    Article Title: A phase I and pharmacokinetics study of intravenous calcitriol in combination with oral dexamethasone and gefitinib in patients with advanced solid tumors

    doi: 10.1007/s00280-009-1000-2

    Figure Lengend Snippet: Treatment schema. On cycle 1, calcitriol ( solid vertical black arrow ) was given on days 1 and 15 and weekly thereafter. Dexamethasone ( broken vertical black arrows ) was given orally 12 h prior, at the time of, and 12 h after to each dose calcitriol. Gefitinib

    Article Snippet: Gefitinib (Iressa) was supplied by AstraZeneca (London, United Kingdom) in 250 mg tablets and was given once daily starting 1 week after the first dose of calcitriol.

    Techniques:

    SKLB188 inhibits the phosphorylation of EGFR and its downstream MEK/Erk and Akt/mTOR pathways in FaDu cells. ( A and B ) Serum-starved FaDu cells were treated with SKLB188 (0–2 μ M ) or Iressa (0 –2 μ M ) for 24 h, followed by stimulation with EGF (50 ng ml −1 ) for 1 h. The cell lysates were subject to western blotting with indicated antibodies.

    Journal: British Journal of Cancer

    Article Title: SKLB188 inhibits the growth of head and neck squamous cell carcinoma by suppressing EGFR signalling

    doi: 10.1038/bjc.2017.298

    Figure Lengend Snippet: SKLB188 inhibits the phosphorylation of EGFR and its downstream MEK/Erk and Akt/mTOR pathways in FaDu cells. ( A and B ) Serum-starved FaDu cells were treated with SKLB188 (0–2 μ M ) or Iressa (0 –2 μ M ) for 24 h, followed by stimulation with EGF (50 ng ml −1 ) for 1 h. The cell lysates were subject to western blotting with indicated antibodies.

    Article Snippet: Reagents SKLB188 was synthesised at the State Key Laboratory of Biotherapy, Sichuan University (Chengdu, China), while Iressa (Gefitinib) was obtained from LC Laboratories (Woburn, MA, USA).

    Techniques: Western Blot

    In vivo anti-tumour activity and mechanism of action of SKLB188. ( A ) Nude mice bearing FaDu tumour cells were treated with SKLB188 or Iressa at the indicated doses or vehicle control alone over 21 days. Points, mean tumour volume or mean body weight; bars, s.d. values. ( B ) At the end of the experiments, the mice were killed and the tumour tissues were dissected and weighed. The data are expressed as the mean±s.d. of groups (6 mice per group). The representative images of isolated tumours are also shown. * P

    Journal: British Journal of Cancer

    Article Title: SKLB188 inhibits the growth of head and neck squamous cell carcinoma by suppressing EGFR signalling

    doi: 10.1038/bjc.2017.298

    Figure Lengend Snippet: In vivo anti-tumour activity and mechanism of action of SKLB188. ( A ) Nude mice bearing FaDu tumour cells were treated with SKLB188 or Iressa at the indicated doses or vehicle control alone over 21 days. Points, mean tumour volume or mean body weight; bars, s.d. values. ( B ) At the end of the experiments, the mice were killed and the tumour tissues were dissected and weighed. The data are expressed as the mean±s.d. of groups (6 mice per group). The representative images of isolated tumours are also shown. * P

    Article Snippet: Reagents SKLB188 was synthesised at the State Key Laboratory of Biotherapy, Sichuan University (Chengdu, China), while Iressa (Gefitinib) was obtained from LC Laboratories (Woburn, MA, USA).

    Techniques: In Vivo, Activity Assay, Mouse Assay, Isolation

    Comparison of Mig-6 expression in lung adenocarcinoma at baseline and after acquiring EGFR-TKI resistance. a–d Representative pictures of Mig-6 by immunohistochemistry in cases 1 and 2. a and c are initial biopsy samples. b and d are matched re-biopsy samples of case 1 and 2. The expression of Mig-6 in re-biopsy samples were significantly higher than those of initial biopsy samples. e and f Analysis of Mig-6 expressions at baseline and after acquiring EGFR-TKI resistance ( n = 26)

    Journal: BMC Cancer

    Article Title: Suppression of Mig-6 overcomes the acquired EGFR-TKI resistance of lung adenocarcinoma

    doi: 10.1186/s12885-020-07057-z

    Figure Lengend Snippet: Comparison of Mig-6 expression in lung adenocarcinoma at baseline and after acquiring EGFR-TKI resistance. a–d Representative pictures of Mig-6 by immunohistochemistry in cases 1 and 2. a and c are initial biopsy samples. b and d are matched re-biopsy samples of case 1 and 2. The expression of Mig-6 in re-biopsy samples were significantly higher than those of initial biopsy samples. e and f Analysis of Mig-6 expressions at baseline and after acquiring EGFR-TKI resistance ( n = 26)

    Article Snippet: The EGFR-TKI gefitinib was obtained from Tocris Bioscience (Iressa, 184,475–35-2; Bristol, UK).

    Techniques: Expressing, Immunohistochemistry

    Graphical summary of Mig-6 function in EGFR-TKI resistant lung adenocarcinoma. Phosphorylated Mig-6 has a crucial oncogenic role in the survival of EGFR-mutant lung adenocarcinoma and targeting Mig-6 may be a promising strategy to overcome EGFR-TKI resistance in lung cancer

    Journal: BMC Cancer

    Article Title: Suppression of Mig-6 overcomes the acquired EGFR-TKI resistance of lung adenocarcinoma

    doi: 10.1186/s12885-020-07057-z

    Figure Lengend Snippet: Graphical summary of Mig-6 function in EGFR-TKI resistant lung adenocarcinoma. Phosphorylated Mig-6 has a crucial oncogenic role in the survival of EGFR-mutant lung adenocarcinoma and targeting Mig-6 may be a promising strategy to overcome EGFR-TKI resistance in lung cancer

    Article Snippet: The EGFR-TKI gefitinib was obtained from Tocris Bioscience (Iressa, 184,475–35-2; Bristol, UK).

    Techniques: Mutagenesis

    Mig-6 is overexpressed in gefitinib-resistant PC9/GR cell lines compared with parental PC9 cell lines. a Mig-6 expression was examined by immunofluorescence staining of endogenous Mig-6 (red) in parental PC9 cells and in PC9/GR cells (Original magnification, × 100 as displayed in the figures), lower panel is optical microscopy images of PC9 and PC9/GR cells (scale bar represents 100 μm). b Western blot experiments for proteins related to epidermal growth factor receptor (EGFR) signaling and the epithelial-mesenchymal transition (EMT) in PC9 and PC9/GR cells. The expressions Mig6, p-EGFR (Tyr1068), p-EGFR (Tyr1045), EGFR, c-MET, p-AKT, AKT, p-ERK, ERK, E-cadherin, Zo-1 and Vimentin were evaluated. β-actin was used as a loading control. c Phos-tag immunoblot analysis of Mig-6 with or without phosphatase. To check the phosphorylated status of Mig-6, the lysates we employed phos-tag SDS-PAGE. The lysate was loaded in 6% Acrylamide 100uM Mn 2+ -Phos-tag™ Acrylamide. To confirm that those slower migrating bands are really phosphorylated species of Mig-6, we treated a part of the lysate with protein phosphatase for 9 h. d RT-PCR analysis. mRNA levels of EGFR, E-cadherin, Vimentin, and c-MET was evaluated in PC9 and PC9/GR. GAPDH was used as an internal control. Uncropped blots were shown in Additional file 1 : Fig. S1

    Journal: BMC Cancer

    Article Title: Suppression of Mig-6 overcomes the acquired EGFR-TKI resistance of lung adenocarcinoma

    doi: 10.1186/s12885-020-07057-z

    Figure Lengend Snippet: Mig-6 is overexpressed in gefitinib-resistant PC9/GR cell lines compared with parental PC9 cell lines. a Mig-6 expression was examined by immunofluorescence staining of endogenous Mig-6 (red) in parental PC9 cells and in PC9/GR cells (Original magnification, × 100 as displayed in the figures), lower panel is optical microscopy images of PC9 and PC9/GR cells (scale bar represents 100 μm). b Western blot experiments for proteins related to epidermal growth factor receptor (EGFR) signaling and the epithelial-mesenchymal transition (EMT) in PC9 and PC9/GR cells. The expressions Mig6, p-EGFR (Tyr1068), p-EGFR (Tyr1045), EGFR, c-MET, p-AKT, AKT, p-ERK, ERK, E-cadherin, Zo-1 and Vimentin were evaluated. β-actin was used as a loading control. c Phos-tag immunoblot analysis of Mig-6 with or without phosphatase. To check the phosphorylated status of Mig-6, the lysates we employed phos-tag SDS-PAGE. The lysate was loaded in 6% Acrylamide 100uM Mn 2+ -Phos-tag™ Acrylamide. To confirm that those slower migrating bands are really phosphorylated species of Mig-6, we treated a part of the lysate with protein phosphatase for 9 h. d RT-PCR analysis. mRNA levels of EGFR, E-cadherin, Vimentin, and c-MET was evaluated in PC9 and PC9/GR. GAPDH was used as an internal control. Uncropped blots were shown in Additional file 1 : Fig. S1

    Article Snippet: The EGFR-TKI gefitinib was obtained from Tocris Bioscience (Iressa, 184,475–35-2; Bristol, UK).

    Techniques: Expressing, Immunofluorescence, Staining, Microscopy, Western Blot, SDS Page, Reverse Transcription Polymerase Chain Reaction

    Mig-6 inhibition restores sensitivity to EGFR-tyrosine kinase inhibitors (TKIs) in PC9/GR cells. a Cell viability was assessed using the CCK-8 assay after transfection with Mig-6 siRNA or scrambled siRNA and treatment with various doses of gefitinib. Data are shown as the mean ± SD; * P

    Journal: BMC Cancer

    Article Title: Suppression of Mig-6 overcomes the acquired EGFR-TKI resistance of lung adenocarcinoma

    doi: 10.1186/s12885-020-07057-z

    Figure Lengend Snippet: Mig-6 inhibition restores sensitivity to EGFR-tyrosine kinase inhibitors (TKIs) in PC9/GR cells. a Cell viability was assessed using the CCK-8 assay after transfection with Mig-6 siRNA or scrambled siRNA and treatment with various doses of gefitinib. Data are shown as the mean ± SD; * P

    Article Snippet: The EGFR-TKI gefitinib was obtained from Tocris Bioscience (Iressa, 184,475–35-2; Bristol, UK).

    Techniques: Inhibition, CCK-8 Assay, Transfection

    Effect of combined treatment of erlotinib or gefitinib with MTE on tumor growth, histological changes in HCC827/ER mice xenografts Tumor volume (A) and weight (B) of HCC827/ER xenografts was assessed. Mice were treated for 3 weeks with vehicle (control), gefitinib or erlotinib (50 mg/kg, p.o.), MTE (5 g/kg, i.p.), or combinations of MTE and one of the 2 other drugs as described in Materials and Methods. The weight of resected tumors was measured after animals were sacrificed. Histological changes (C) were detected by HE staining (200 ×) and immunohistochemistry staining to compare tumor growth (PCNA), cell apoptosis (TUNEL), tumor angiogenesis (CD105, VEGF), and EMT makers (E-cadherin, vimentin) between various treatment groups. Data are presented as the mean ± SE from mice in each group (n = 8). *p

    Journal: Oncotarget

    Article Title: Marsdenia tenacissima extract overcomes Axl- and Met-mediated erlotinib and gefitinib cross-resistance in non-small cell lung cancer cells

    doi: 10.18632/oncotarget.18137

    Figure Lengend Snippet: Effect of combined treatment of erlotinib or gefitinib with MTE on tumor growth, histological changes in HCC827/ER mice xenografts Tumor volume (A) and weight (B) of HCC827/ER xenografts was assessed. Mice were treated for 3 weeks with vehicle (control), gefitinib or erlotinib (50 mg/kg, p.o.), MTE (5 g/kg, i.p.), or combinations of MTE and one of the 2 other drugs as described in Materials and Methods. The weight of resected tumors was measured after animals were sacrificed. Histological changes (C) were detected by HE staining (200 ×) and immunohistochemistry staining to compare tumor growth (PCNA), cell apoptosis (TUNEL), tumor angiogenesis (CD105, VEGF), and EMT makers (E-cadherin, vimentin) between various treatment groups. Data are presented as the mean ± SE from mice in each group (n = 8). *p

    Article Snippet: Drugs and reagents Gefitinib (Iressa) was obtained from AstraZeneca (Cheshire, UK) and erlotinib (Tarceva) was purchased from Roche Pharma (Schweiz, UK).

    Techniques: Mouse Assay, Staining, Immunohistochemistry, TUNEL Assay

    Effects of combined treatment of erlotinib or gefitinib with MTE on cell viability and apoptosis in HCC827/ER cells (A) The viability of parental HCC827 cells and resistant HCC827/ER cells after treatment with the indicated concentration of MTE for 72h. HCC827/ER cells were treated with erlotinib (B) , gefitinib (C) , or three different combinations of 8 mg/ml MTE and one of the other 2 drugs for 72 hours. Results were expressed as the percentage of living cells compared to the control, and error bars indicated SD of three independent measurements.* p

    Journal: Oncotarget

    Article Title: Marsdenia tenacissima extract overcomes Axl- and Met-mediated erlotinib and gefitinib cross-resistance in non-small cell lung cancer cells

    doi: 10.18632/oncotarget.18137

    Figure Lengend Snippet: Effects of combined treatment of erlotinib or gefitinib with MTE on cell viability and apoptosis in HCC827/ER cells (A) The viability of parental HCC827 cells and resistant HCC827/ER cells after treatment with the indicated concentration of MTE for 72h. HCC827/ER cells were treated with erlotinib (B) , gefitinib (C) , or three different combinations of 8 mg/ml MTE and one of the other 2 drugs for 72 hours. Results were expressed as the percentage of living cells compared to the control, and error bars indicated SD of three independent measurements.* p

    Article Snippet: Drugs and reagents Gefitinib (Iressa) was obtained from AstraZeneca (Cheshire, UK) and erlotinib (Tarceva) was purchased from Roche Pharma (Schweiz, UK).

    Techniques: Concentration Assay

    Effect of combined treatment of erlotinib or gefitinib with MTE on EGFR-related downstream molecules in HCC827/ER cells Cells were treated with 1 μM erlotinib or gefitinib alone or in combination with 8 mg/ml MTE (M→M+E for erlotinib, M→M+G for gefitinib) for 24 hours. Cells were stimulated with 10 ng/ml EGF for 15 min before harvest. Cell lysates were collected and subjected to SDS-PAGE and Western blotting, analyzed with phospho-specific antibodies to PI3K/Akt/mTOR, ERK1/2 (A for gefitinib, C for erlotinib), EGFR and c-Met ( E for gefitinib, G for erlotinib). (B and D) are quantification of (A and C) , (F and H) are quantification of (E and G) , respectively. Each bar represents mean ± SD of three separate experiments. * p

    Journal: Oncotarget

    Article Title: Marsdenia tenacissima extract overcomes Axl- and Met-mediated erlotinib and gefitinib cross-resistance in non-small cell lung cancer cells

    doi: 10.18632/oncotarget.18137

    Figure Lengend Snippet: Effect of combined treatment of erlotinib or gefitinib with MTE on EGFR-related downstream molecules in HCC827/ER cells Cells were treated with 1 μM erlotinib or gefitinib alone or in combination with 8 mg/ml MTE (M→M+E for erlotinib, M→M+G for gefitinib) for 24 hours. Cells were stimulated with 10 ng/ml EGF for 15 min before harvest. Cell lysates were collected and subjected to SDS-PAGE and Western blotting, analyzed with phospho-specific antibodies to PI3K/Akt/mTOR, ERK1/2 (A for gefitinib, C for erlotinib), EGFR and c-Met ( E for gefitinib, G for erlotinib). (B and D) are quantification of (A and C) , (F and H) are quantification of (E and G) , respectively. Each bar represents mean ± SD of three separate experiments. * p

    Article Snippet: Drugs and reagents Gefitinib (Iressa) was obtained from AstraZeneca (Cheshire, UK) and erlotinib (Tarceva) was purchased from Roche Pharma (Schweiz, UK).

    Techniques: SDS Page, Western Blot

    Effect of combined treatment of erlotinib or gefitinib with MTE on EGFR bypass signaling in HCC827/ER cells Cells were treated with 1 μM erlotinib and gefitinib alone or in combination with 8 mg/ml MTE (M→M+E for erlotinib, M→M+G fore gefitinib) for 24 hours. To determine the expression profile of p-Met, cells were stimulated with 10 ng/ml HGF for 5 min before harvest (A-D). Cells harvested for detection of other bypass signals were not stimulated with HGF (E-H). Cell lysates were collected and subjected to SDS-PAGE and Western blotting, analyzed with phospho-specific antibodies to p-Met ( A for gefitinib, C for erlotinib), p-Axl, E-cadherin, N-cadherin, vimentin and p-P70S6K ( E for gefitinib, G for erlotinib). (B and D) are quantification of the results in (A and C) , and (F and H) are quantification of (E and G). Each bar represents mean ± SD of three separate experiments. * p

    Journal: Oncotarget

    Article Title: Marsdenia tenacissima extract overcomes Axl- and Met-mediated erlotinib and gefitinib cross-resistance in non-small cell lung cancer cells

    doi: 10.18632/oncotarget.18137

    Figure Lengend Snippet: Effect of combined treatment of erlotinib or gefitinib with MTE on EGFR bypass signaling in HCC827/ER cells Cells were treated with 1 μM erlotinib and gefitinib alone or in combination with 8 mg/ml MTE (M→M+E for erlotinib, M→M+G fore gefitinib) for 24 hours. To determine the expression profile of p-Met, cells were stimulated with 10 ng/ml HGF for 5 min before harvest (A-D). Cells harvested for detection of other bypass signals were not stimulated with HGF (E-H). Cell lysates were collected and subjected to SDS-PAGE and Western blotting, analyzed with phospho-specific antibodies to p-Met ( A for gefitinib, C for erlotinib), p-Axl, E-cadherin, N-cadherin, vimentin and p-P70S6K ( E for gefitinib, G for erlotinib). (B and D) are quantification of the results in (A and C) , and (F and H) are quantification of (E and G). Each bar represents mean ± SD of three separate experiments. * p

    Article Snippet: Drugs and reagents Gefitinib (Iressa) was obtained from AstraZeneca (Cheshire, UK) and erlotinib (Tarceva) was purchased from Roche Pharma (Schweiz, UK).

    Techniques: Expressing, SDS Page, Western Blot

    Effects of MTE and erlotinib or gefitinib combination treatment on EGFR downstream pathways and bypass signaling pathways in xenograft tumors Mice were treated as described previously. The resected tumors were lysed and total protein collected to measure EGFR related signaling molecules by Western blotting. The combined effects of MTE and erlotinib or gefitinib were evaluated through detecting the expression of EGFR downstream PI3K/Akt/mTOR and ERK ( A for gefitinib, C for erlotinib), activation of bypass pathway markers c-Met and Axl ( E for gefitinib, G for erlotinib). (B and D) are quantification of results found in (A and C) , and (F and H) are quantification of (E and G). Each bar represents mean ± SD of three separate experiments. * p

    Journal: Oncotarget

    Article Title: Marsdenia tenacissima extract overcomes Axl- and Met-mediated erlotinib and gefitinib cross-resistance in non-small cell lung cancer cells

    doi: 10.18632/oncotarget.18137

    Figure Lengend Snippet: Effects of MTE and erlotinib or gefitinib combination treatment on EGFR downstream pathways and bypass signaling pathways in xenograft tumors Mice were treated as described previously. The resected tumors were lysed and total protein collected to measure EGFR related signaling molecules by Western blotting. The combined effects of MTE and erlotinib or gefitinib were evaluated through detecting the expression of EGFR downstream PI3K/Akt/mTOR and ERK ( A for gefitinib, C for erlotinib), activation of bypass pathway markers c-Met and Axl ( E for gefitinib, G for erlotinib). (B and D) are quantification of results found in (A and C) , and (F and H) are quantification of (E and G). Each bar represents mean ± SD of three separate experiments. * p

    Article Snippet: Drugs and reagents Gefitinib (Iressa) was obtained from AstraZeneca (Cheshire, UK) and erlotinib (Tarceva) was purchased from Roche Pharma (Schweiz, UK).

    Techniques: Mouse Assay, Western Blot, Expressing, Activation Assay

    Cytotoxicity of EGFR-TKIs and molecular profiles in parental HCC827 and resistant cell line HCC827/ER Cells were treated with the indicated concentrations of erlotinib (A) and gefitinib (B) for 72 h in medium containing 1% FBS. Cell viability was determined using an MTT assay, and IC 50 values were calculated using Graphpad Prism software 5.0. Results were expressed as the percentage of living cells compared to the control, error bars indicate SD of three independent measurements. * p

    Journal: Oncotarget

    Article Title: Marsdenia tenacissima extract overcomes Axl- and Met-mediated erlotinib and gefitinib cross-resistance in non-small cell lung cancer cells

    doi: 10.18632/oncotarget.18137

    Figure Lengend Snippet: Cytotoxicity of EGFR-TKIs and molecular profiles in parental HCC827 and resistant cell line HCC827/ER Cells were treated with the indicated concentrations of erlotinib (A) and gefitinib (B) for 72 h in medium containing 1% FBS. Cell viability was determined using an MTT assay, and IC 50 values were calculated using Graphpad Prism software 5.0. Results were expressed as the percentage of living cells compared to the control, error bars indicate SD of three independent measurements. * p

    Article Snippet: Drugs and reagents Gefitinib (Iressa) was obtained from AstraZeneca (Cheshire, UK) and erlotinib (Tarceva) was purchased from Roche Pharma (Schweiz, UK).

    Techniques: MTT Assay, Software

    Lapatinib attenuates HER2CA GH3 tumor growth and hormone secretion in vivo more than gefitinib. A, GH3 cells stably expressing HER2CA (3 × 10 6 cells per rat, 0.2 ml with matrigel) or pcDNA3 were inoculated sc in WF rats (4–5 wk of age).

    Journal: Molecular Endocrinology

    Article Title: HER2/ErbB2 Receptor Signaling in Rat and Human Prolactinoma Cells: Strategy for Targeted Prolactinoma Therapy

    doi: 10.1210/me.2010-0353

    Figure Lengend Snippet: Lapatinib attenuates HER2CA GH3 tumor growth and hormone secretion in vivo more than gefitinib. A, GH3 cells stably expressing HER2CA (3 × 10 6 cells per rat, 0.2 ml with matrigel) or pcDNA3 were inoculated sc in WF rats (4–5 wk of age).

    Article Snippet: Lapatinib (Tykarb) was from LC Laboratories (Woburn, MA), and gefitinib (Iressa) was purchased from Biaffin GmbH & Co. (Kassel, Germany).

    Techniques: In Vivo, Stable Transfection, Expressing

    Lapatinib attenuates HER2 signaling and suppresses PRL more than gefitinib. A, GH3 cells stably expressing HER2CA were pretreated with gefitinib or lapatinib (0.1–10 μ m ) for 45 min before induction with EGF (5 n m ) for 10 min, and Western

    Journal: Molecular Endocrinology

    Article Title: HER2/ErbB2 Receptor Signaling in Rat and Human Prolactinoma Cells: Strategy for Targeted Prolactinoma Therapy

    doi: 10.1210/me.2010-0353

    Figure Lengend Snippet: Lapatinib attenuates HER2 signaling and suppresses PRL more than gefitinib. A, GH3 cells stably expressing HER2CA were pretreated with gefitinib or lapatinib (0.1–10 μ m ) for 45 min before induction with EGF (5 n m ) for 10 min, and Western

    Article Snippet: Lapatinib (Tykarb) was from LC Laboratories (Woburn, MA), and gefitinib (Iressa) was purchased from Biaffin GmbH & Co. (Kassel, Germany).

    Techniques: Stable Transfection, Expressing, Western Blot

    Dose-dependent effects of gefitinib or lapatinib on HER2CA GH3 proliferation and apoptosis. A, GH3 cells stably expressing HER2CA were treated with gefitinib or lapatinib (0.1–10 μ m ) for 24 h, and cells were counted. B, Stably transduced

    Journal: Molecular Endocrinology

    Article Title: HER2/ErbB2 Receptor Signaling in Rat and Human Prolactinoma Cells: Strategy for Targeted Prolactinoma Therapy

    doi: 10.1210/me.2010-0353

    Figure Lengend Snippet: Dose-dependent effects of gefitinib or lapatinib on HER2CA GH3 proliferation and apoptosis. A, GH3 cells stably expressing HER2CA were treated with gefitinib or lapatinib (0.1–10 μ m ) for 24 h, and cells were counted. B, Stably transduced

    Article Snippet: Lapatinib (Tykarb) was from LC Laboratories (Woburn, MA), and gefitinib (Iressa) was purchased from Biaffin GmbH & Co. (Kassel, Germany).

    Techniques: Stable Transfection, Expressing

    Sprr2d and Slpi expression in infundibula of K5cre-CMVcaNrf2 mice. A Average number of BrdU-positive cells in infundibula (INF) of tg/wt and tg/tg mice ( N = 4). B,C Average thickness of infundibula and number of PCNA-positive cells in infundibula of P32 tg/wt and tg/tg mice injected with vehicle or Gefitinib ( N = 4/5). D Upper panel: immunohistochemistry of Sprr2 on longitudinal P10 tg/wt and tg/tg back skin sections. Note staining of Sprr2 in differentiated infundibular keratinocytes in tg/tg mice. Scale bar: 25 μm. Middle panel: toluidine blue staining for detection of mast cells on longitudinal back skin sections of P32 tg/wt and tg/tg mice. Note increase in number of mast cells in tg/tg mice and assembly of mast cells along the interfollicular epidermis and upper part of the hair follicles. Scale bar: 100 μm. Lower panel: immunohistochemistry of Slpi on longitudinal P10 tg/wt and tg/tg back skin sections. Inset in lower panel shows immunohistochemistry without primary antibody. The dashed line marks the basement membrane of the interfollicular epidermis. Note staining of Slpi (indicated by arrow) in differentiated infundibular keratinocytes and stratum corneum in tg/tg mice. Scale bar: 25 μm. E Electron microscopy of infundibular stratum corneum of P32 tg/wt and tg/tg mice. Corneocyte layers in lower panel were numbered from basal to distal (C1–C3). Arrows point to corneodesmosomes. Note delayed corneodesmosome degradation in tg/tg mice. C, corneocyte. F Working model: Nrf2 activation leads to upregulation of Slpi, Sprr2d, and Epgn in hair follicle infundibula. Slpi upregulation promotes inhibition of CE protease activity. Consequently, corneodesmosome cleavage is reduced, leading to decreased desquamation and thereby to hyperkeratosis of infundibula. Sprr2d upregulation reduces epidermal barrier functionality, resulting in increased inflammation and consequently stimulation of proliferation. Epgn and other epidermal growth factor (EGF) family members stimulate proliferation of infundibular keratinocytes via EGFR signaling. Together, this results in acanthosis of hair follicle infundibula. Data information: Values are shown as the mean with s.d.

    Journal: EMBO Molecular Medicine

    Article Title: Activation of Nrf2 in keratinocytes causes chloracne (MADISH)-like skin disease in mice

    doi: 10.1002/emmm.201303281

    Figure Lengend Snippet: Sprr2d and Slpi expression in infundibula of K5cre-CMVcaNrf2 mice. A Average number of BrdU-positive cells in infundibula (INF) of tg/wt and tg/tg mice ( N = 4). B,C Average thickness of infundibula and number of PCNA-positive cells in infundibula of P32 tg/wt and tg/tg mice injected with vehicle or Gefitinib ( N = 4/5). D Upper panel: immunohistochemistry of Sprr2 on longitudinal P10 tg/wt and tg/tg back skin sections. Note staining of Sprr2 in differentiated infundibular keratinocytes in tg/tg mice. Scale bar: 25 μm. Middle panel: toluidine blue staining for detection of mast cells on longitudinal back skin sections of P32 tg/wt and tg/tg mice. Note increase in number of mast cells in tg/tg mice and assembly of mast cells along the interfollicular epidermis and upper part of the hair follicles. Scale bar: 100 μm. Lower panel: immunohistochemistry of Slpi on longitudinal P10 tg/wt and tg/tg back skin sections. Inset in lower panel shows immunohistochemistry without primary antibody. The dashed line marks the basement membrane of the interfollicular epidermis. Note staining of Slpi (indicated by arrow) in differentiated infundibular keratinocytes and stratum corneum in tg/tg mice. Scale bar: 25 μm. E Electron microscopy of infundibular stratum corneum of P32 tg/wt and tg/tg mice. Corneocyte layers in lower panel were numbered from basal to distal (C1–C3). Arrows point to corneodesmosomes. Note delayed corneodesmosome degradation in tg/tg mice. C, corneocyte. F Working model: Nrf2 activation leads to upregulation of Slpi, Sprr2d, and Epgn in hair follicle infundibula. Slpi upregulation promotes inhibition of CE protease activity. Consequently, corneodesmosome cleavage is reduced, leading to decreased desquamation and thereby to hyperkeratosis of infundibula. Sprr2d upregulation reduces epidermal barrier functionality, resulting in increased inflammation and consequently stimulation of proliferation. Epgn and other epidermal growth factor (EGF) family members stimulate proliferation of infundibular keratinocytes via EGFR signaling. Together, this results in acanthosis of hair follicle infundibula. Data information: Values are shown as the mean with s.d.

    Article Snippet: EGFR signaling was inhibited by intraperitoneal injection of P18 K5cre-CMVcaNrf2 and control mice with 70 μg Gefitinib (Iressa) (Santa Cruz) per gram body weight once a day for 14 consecutive days.

    Techniques: Expressing, Mouse Assay, Injection, Immunohistochemistry, Staining, Electron Microscopy, Activation Assay, Inhibition, Activity Assay

    Regulation of Epgn by Nrf2. A Average SG area of P32 tg/wt and tg/tg mice injected with vehicle or Gefitinib. Note reduction in SG area upon Gefitinib treatment of tg/wt mice ( N = 6/5, ** P = 0.0025) and tg/tg mice ( N = 4, * P = 0.0286). B, C qRT-PCR of Adph , Pparg Mc5r (B), and Scd1 , 2 , 3 , 4 (C) relative to Gapdh using RNA of back skin from wt and EPGN -tg mice ( N = 3). D, E qRT-PCR of Epgn using RNAs of murine primary keratinocytes of tg/wt and tg/tg mice ( N = 3) (D), or immortalized keratinocytes of wt and Nrf2ko mice treated with vehicle or sulforaphane (E) ( N = 4, * P = 0.0286). F Localization of PCR-amplified non-specific region (ns), promoter region (prom), and antioxidant response elements (AREs) in the murine Nqo1 and Epgn genes. G Chromatin immunoprecipitation (ChIP) using lysates from back and tail skin of control mice. Note the enhanced binding of Nrf2 to the Epgn ARE compared with the ns region ( Nqo1 ns, N = 6, Nqo1 ARE, N = 6, ** P = 0.0045; Epgn ARE, N = 4, * P = 0.012). H, I ChIP using back and tail skin lysates of tg/wt (H) and tg/tg mice (I) with an H3K4me2 antibody. Regions void of annotated ORFs (non-coding, nc DNA), Epgn prom, and Epgn ARE were amplified by qRT-PCR. Note the increase in histone H3 dimethylation at the Epgn ARE compared with ncDNA in tg/tg mice ( N = 2/3). J Working model: Nrf2 activation leads to increased Epgn expression, which activates EGFR signaling. This enhances proliferation of sebocyte progenitors, leading to sebaceous gland hyperplasia. Data information: Values are shown as the mean with s.d. All P -values were calculated by Mann–Whitney U -test.

    Journal: EMBO Molecular Medicine

    Article Title: Activation of Nrf2 in keratinocytes causes chloracne (MADISH)-like skin disease in mice

    doi: 10.1002/emmm.201303281

    Figure Lengend Snippet: Regulation of Epgn by Nrf2. A Average SG area of P32 tg/wt and tg/tg mice injected with vehicle or Gefitinib. Note reduction in SG area upon Gefitinib treatment of tg/wt mice ( N = 6/5, ** P = 0.0025) and tg/tg mice ( N = 4, * P = 0.0286). B, C qRT-PCR of Adph , Pparg Mc5r (B), and Scd1 , 2 , 3 , 4 (C) relative to Gapdh using RNA of back skin from wt and EPGN -tg mice ( N = 3). D, E qRT-PCR of Epgn using RNAs of murine primary keratinocytes of tg/wt and tg/tg mice ( N = 3) (D), or immortalized keratinocytes of wt and Nrf2ko mice treated with vehicle or sulforaphane (E) ( N = 4, * P = 0.0286). F Localization of PCR-amplified non-specific region (ns), promoter region (prom), and antioxidant response elements (AREs) in the murine Nqo1 and Epgn genes. G Chromatin immunoprecipitation (ChIP) using lysates from back and tail skin of control mice. Note the enhanced binding of Nrf2 to the Epgn ARE compared with the ns region ( Nqo1 ns, N = 6, Nqo1 ARE, N = 6, ** P = 0.0045; Epgn ARE, N = 4, * P = 0.012). H, I ChIP using back and tail skin lysates of tg/wt (H) and tg/tg mice (I) with an H3K4me2 antibody. Regions void of annotated ORFs (non-coding, nc DNA), Epgn prom, and Epgn ARE were amplified by qRT-PCR. Note the increase in histone H3 dimethylation at the Epgn ARE compared with ncDNA in tg/tg mice ( N = 2/3). J Working model: Nrf2 activation leads to increased Epgn expression, which activates EGFR signaling. This enhances proliferation of sebocyte progenitors, leading to sebaceous gland hyperplasia. Data information: Values are shown as the mean with s.d. All P -values were calculated by Mann–Whitney U -test.

    Article Snippet: EGFR signaling was inhibited by intraperitoneal injection of P18 K5cre-CMVcaNrf2 and control mice with 70 μg Gefitinib (Iressa) (Santa Cruz) per gram body weight once a day for 14 consecutive days.

    Techniques: Mouse Assay, Injection, Quantitative RT-PCR, Polymerase Chain Reaction, Amplification, Chromatin Immunoprecipitation, Binding Assay, Activation Assay, Expressing, MANN-WHITNEY