Journal: Cell Death & Disease

Article Title: Targeting the COX2/MET/TOPK signaling axis induces apoptosis in gefitinib-resistant NSCLC cells

doi: 10.1038/s41419-019-2020-4

Figure Lengend Snippet: a The levels of PGs were measured in the supernatants from HCC827 and HCC827GR cells. b Measurement of TXA 2 and PGE 2 in supernatants from shCOX2 HCC827GR cells (left), HCC827 cells stably expressing COX2 (middle), or HCC827GR cells treated with celecoxib for 48 h at different doses (right). c HCC827 cells were treated with U46619 for 48 h at different doses (left); HCC827GR cells were treated with different doses of Seratrodast (middle) or Ozarel HCl (right) for 48 h. d HCC827 cells stably expressing COX2 (left) or HCC827GR cells (right) were treated with SR11302 for 48 h. e The stable shCOX2 HCC827GR cells were treated with gefitinib (1 μΜ) for 6 h (left); HCC827GR cells were treated with different doses of celecoxib for 48 h (right). Those samples were resolved by SDS-PAGE and visualized by western blotting. Data are representatives of results from triplicate experiments. * P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: Gefitinib was purchased from Gold Biotechnology (St. Louis, MO).

Techniques: Stable Transfection, Expressing, SDS Page, Western Blot

Journal: Cell Death & Disease

Article Title: Targeting the COX2/MET/TOPK signaling axis induces apoptosis in gefitinib-resistant NSCLC cells

doi: 10.1038/s41419-019-2020-4

Figure Lengend Snippet: a Western blotting analysis in different lung cancer cells. b HCC827 and HCC827GR cells were exposed to increasing concentrations of gefitinib for 6 h. The samples were separated by SDS-PAGE and visualized by western blotting. c IHC analysis of MET and TOPK expression in TMA. Magnifications, ×40, ×100, ×200. d The correlation between MET and TOPK expression in EGFR -activating mutated NSCLC was analyzed. Data are representative of results from triplicate experiments. *** P < 0.001; R = 0.1801

Article Snippet: Gefitinib was purchased from Gold Biotechnology (St. Louis, MO).

Techniques: Western Blot, SDS Page, Expressing

Journal: Cell Death & Disease

Article Title: Targeting the COX2/MET/TOPK signaling axis induces apoptosis in gefitinib-resistant NSCLC cells

doi: 10.1038/s41419-019-2020-4

Figure Lengend Snippet: a Active MET phosphorylated inactive TOPK-WT in vitro in the presence of [γ- 32 P] ATP as visualized by autoradiography. b Potential phosphorylated Tyr sites of TOPK were predicted by the NetPhos 3.0 software program. c Synthesized peptides containing potential Tyr b sites were used as substrates in an in vitro kinase assay with active MET in the presence of [γ- 32 P] ATP and the results were visualized by autoradiography. d Active MET phosphorylated inactive TOPK-WT or 74F in vitro in the presence of ATP. Then, the samples were analyzed by western blotting. e MET promoted the phosphorylation of TOPK in HEK293T cells induced by HGF after transfection with His-MET in a dose-dependent manner (HGF 40 ng/ml; 15 min). f His-MET was cotransfected in HEK293T cells with wild-type TOPK (HA-TOPK WT) or Y74-mutated TOPK (HA-TOPK 74F). g TOPK bound with endogenous MET of HCC827GR cells by IP assay. h The stable shMock and shMET in HCC827GR cells were treated with gefitinib (1 μΜ) for 6 h. i HCC827 cells stably overexpressing Mock, HA-TOPK WT, and HA-TOPK 74F were cultured and analyzed by western blotting. The above samples were collected and analyzed by western blotting. Data are representative of results from triplicate experiments

Article Snippet: Gefitinib was purchased from Gold Biotechnology (St. Louis, MO).

Techniques: In Vitro, Autoradiography, Software, Synthesized, Kinase Assay, Western Blot, Transfection, Stable Transfection, Cell Culture

Journal: Cell Death & Disease

Article Title: Targeting the COX2/MET/TOPK signaling axis induces apoptosis in gefitinib-resistant NSCLC cells

doi: 10.1038/s41419-019-2020-4

Figure Lengend Snippet: a , b COX2 (left) or TOPK (right) gene silencing in HCC827GR cells promoted cell apoptosis with gefitinib (1 μΜ) treatment for 24 h. Total and cleaved PARP were detected by western blotting analysis a . Apoptotic cells were detected by Annexin V–FITC and PI double staining and analyzed by flow cytometry b . c , d Stably expressing COX2 (left) or HA-TOPK WT (right) in HCC827 cells inhibited cell apoptosis; stably expressing HA-TOPK 74F (right) promoted cell apoptosis. The cells were treated with gefitinib (1 μΜ) for 24 h. Total and cleaved PARP and apoptotic cells were detected as same as a , b . e , f Transfectants of HCC827 (Mock), (COX2), (TOPK-WT) or (74F) were compared for colony formation by soft agar assay with gefitinib treatment (0 μΜ, 0.5 μΜ, 1 μΜ, 2 μΜ). Scale bar: 500 μm. ** P < 0.01, *** P < 0.001. Data are representative of results from triplicate experiments

Article Snippet: Gefitinib was purchased from Gold Biotechnology (St. Louis, MO).

Techniques: Western Blot, Double Staining, Flow Cytometry, Stable Transfection, Expressing, Soft Agar Assay

Journal: Cell Death & Disease

Article Title: Targeting the COX2/MET/TOPK signaling axis induces apoptosis in gefitinib-resistant NSCLC cells

doi: 10.1038/s41419-019-2020-4

Figure Lengend Snippet: a Combined effects were measured using CI values. Quantification of the potency of the different triple combinations using the Chou–Talalay method as described in the Materials and Methods section. Fa: Fraction affected. CI values < 1.0 correspond to synergistic inhibition effects, and the CI of gefitinib (1 μΜ), celecoxib (20 μΜ), and pantoprazole (100 μΜ) in HCC827GR was 0.495. b HCC827GR cells were treated with gefitinib (1 μΜ), celecoxib (20 μΜ), and pantoprazole (100 μΜ) alone, with the three distinct combinations of two inhibitors and with the triple combination for 24, 48, or 72 h. Then, the effects on cell growth were measured by MTT assay. Bars represent the SD of experiments repeated in triplicate. *** P < 0.001 for the triple combination of drugs vs. either control or drug alone or other two drugs. c HCC827GR cells were treated with the same drugs as b for 24 h and the samples were analyzed by western blotting. d The colonies in soft agar were compared in HCC827GR cells treated with gefitinib, celecoxib, or pantoprazole. Scale bar: 500 μm. *** P < 0.001 for the triple combination of drugs vs. either control or drug alone or other two drugs. e , f Mice were treated by intraperitoneal injection with vehicle, gefitinib, or gefitinib plus celecoxib plus pantoprazole for 19 days. Significant differences were determined by factorial analysis of variance (* P < 0.05, ** P < 0.01, *** P < 0.001 vs. either vehicle or gefitinib alone). Representative photographs show the external appearance of tumors. g PFS analysis between COX2-, MET-, and TOPK-positive and -negative patients with EGFR -activating mutations. * P < 0.05. Data are represented as the means ± SD of triplicate experiments. h Schematic diagram showing the mechanism of COX2/MET/TOPK signaling in gefitinib-resistant cells

Article Snippet: Gefitinib was purchased from Gold Biotechnology (St. Louis, MO).

Techniques: Inhibition, MTT Assay, Western Blot, Injection

Journal: Oncotarget

Article Title: Shp2 regulates migratory behavior and response to EGFR-TKIs through ERK1/2 pathway activation in non-small cell lung cancer cells

doi: 10.18632/oncotarget.20249

Figure Lengend Snippet: (A) , (B) Shp2 knockdown decreased the proliferation of H1975 cells (A) and H292 cells (B). Lung cancer cells were transfected with Shp2 or control siRNA for 72 h in RPMI 1640 medium containing 0.5% FBS. Cell proliferation was measured using the CCK8 assay at 72 h after transfection, and the results are expressed as means ± SD (N = 4); * P < 0.05 versus control. (C) , (D) Shp2 knockdown enhanced gefitinib sensitivity in NSCLC cells. (E) , (F) Shp2 overexpression decreased gefitinib sensitivity in NSCLC cells. H1975 and H292 cells transfected with Shp2/control siRNAs or Shp2 WT /vector were treated with 0–10 μM gefitinib for 3 days, and then cell proliferation was measured using the CCK8 assay. The cell-viability percentage was calculated by adjusting the DMSO control group to 100%. Points and error bars represent the mean and SD values from 3 independent experiments. Black points: control siRNAs/vector; Gray points: Shp2-siRNAs/Shp2 WT .

Article Snippet: The ERK1/2 pharmacological inhibitor U0126 was purchased from Calbiochem (Merck, Darmstadt, Germany) and the EGFR-TKIs gefitinib (Iressa) was from Tocris Bioscience (Bristol, UK).

Techniques: Knockdown, Transfection, Control, CCK-8 Assay, Over Expression, Plasmid Preparation

Journal: Oncotarget

Article Title: Shp2 regulates migratory behavior and response to EGFR-TKIs through ERK1/2 pathway activation in non-small cell lung cancer cells

doi: 10.18632/oncotarget.20249

Figure Lengend Snippet: (A) Comparison of gefitinib sensitivity of H292 cells transfected with vector or Shp2 WT and treated with DMSO or U0126; cell survival was measured using the CCK8 assay. (B) U0126 treatment abrogated Shp2 WT -induced upregulation of c-Myc and fibronectin expression and downregulation of E-cadherin expression in H292 cells. Equal amounts of lysates from cells transfected with vector/Shp2 WT were immunoblotted with the indicated antibodies. (C) , (D) U0126 inhibits H292 cell migration; cells treated with DMSO or U0126 were used in wound-healing assays (C) and Transwell-migration assays (D). * P < 0.05 versus controls.

Article Snippet: The ERK1/2 pharmacological inhibitor U0126 was purchased from Calbiochem (Merck, Darmstadt, Germany) and the EGFR-TKIs gefitinib (Iressa) was from Tocris Bioscience (Bristol, UK).

Techniques: Comparison, Transfection, Plasmid Preparation, CCK-8 Assay, Expressing, Migration

Journal: bioRxiv

Article Title: Candida albicans stimulates the formation of a multi-receptor complex that mediates epithelial cell invasion during oropharyngeal infection

doi: 10.1101/2023.02.23.529756

Figure Lengend Snippet: (A-C) Immunoblot analysis showing the effects of the EGFR inhibitor gefitinib and the c-Met inhibitor SGX523 on C. albicans -induced phosphorylation of c-Met and EGFR in oral epithelial cells after 20 min of infection. Representative immunoblot (A), densitometric analysis of 4 immunoblots showing the phosphorylation of EGFR (B) and c-Met (C). Data are mean ± SD. (D) Effects of SGX523 and gefitinib on the endocytosis of C. albicans by oral epithelial cells. (E and F) Endocytosis of C. albicans by NIH/3T3 cells that expressed human c-Met (E) or human c-Met, human EGFR, and human Her2 (F). (G-I) Knockdown of E-cadherin by siRNA inhibits the phosphorylation of c-Met and EGFR in oral epithelial cells infected with C. albicans . Representative immunoblot (G). Densitometric analysis of 5 immunoblots showing the phosphorylation of c-Met (H) and EGFR (I). Results are mean ± SD. J and K) Effects of inhibiting c-Met (J) and EGFR (K) in combination with siRNA knockdown of E-cadherin on the endocytosis of C. albicans by oral epithelial cells. Results in (D-F, J, and K) are the mean ± SD of three experiments, each performed in triplicate. * p < 05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, ns; not significant (one-way ANOVA with Sidak’s multiple comparisons test [B-D, H-K] or two-tailed Student’s t test [E and F]).

Article Snippet: Gefitinib (#S1025, Selleck Chem) was dissolved in DMSO at concentration of 100 mM, and diluted further to 1 μM in KSF medium (#17005042, Thermo Fisher Scientific) without supplements for final use.

Techniques: Western Blot, Phospho-proteomics, Infection, Knockdown, Two Tailed Test

Journal: bioRxiv

Article Title: Candida albicans stimulates the formation of a multi-receptor complex that mediates epithelial cell invasion during oropharyngeal infection

doi: 10.1101/2023.02.23.529756

Figure Lengend Snippet: (A) Hyr1 and Als3 are required for C. albicans to resist killing by human neutrophils. Neutrophils were infected with the indicated C. albicans strains constructed in the SC5314 strain background. Results are mean ± SD of neutrophils from 3 donors, tested in triplicate. (B) The hyr1 Δ/Δ mutant is resistant to neutrophil killing, even in the absence of c-Met. Killing of wild-type and hyr1 Δ/Δ mutant strains in the SN250 strain background by neutrophils from Mrp8; Met fl/fl mice, which have a neutrophil-specific deletion in c-Met, and their littermates. Results are mean ± SD of neutrophils from 2 experiments performed in triplicate. (C and D) Oral fungal burden of immunocompetent mice infected with the indicated C. albicans strains for 1 (C) and 2 (D) days. (E and F) Effects of phagocyte depletion on susceptibility to OPC, as determined by oral fungal burden after 1 (E) and 2 (F) days of infection. (G-H) Oral fungal burden after 5 d of infection of mice that had been immunosuppressed with cortisone acetate and treated with gefitinib and/or SGX523 (G). Histopathology of the tongues (H). Scale bar 200 µm. Data are the mean ± SD combined results from 2 independent experiments, each using 4 mice per C. albicans strain (C-F) or 8 mice per condition in one experiment and 5 mice per condition in the other (G). ns, not significant; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 (one-way ANOVA with Sidak’s multiple comparisons test)

Article Snippet: Gefitinib (#S1025, Selleck Chem) was dissolved in DMSO at concentration of 100 mM, and diluted further to 1 μM in KSF medium (#17005042, Thermo Fisher Scientific) without supplements for final use.

Techniques: Infection, Construct, Mutagenesis, Histopathology

Journal: Cancer research and treatment

Article Title: miR-4487 Enhances Gefitinib-Mediated Ubiquitination and Autophagic Degradation of EGFR in Non-Small Cell Lung Cancer Cells by Targeting USP37.

doi: 10.4143/crt.2021.622

Figure Lengend Snippet: Fig. 1. Effects of long-term treatment of gefitinib on non–small cell lung cancer cells. Different cell lines were incubated with gefitinib (1 μM) for the indicated time. (A) Endogenous levels of epidermal growth factor receptor (EGFR) and cleavage of poly(ADP-ribose) poly merase-1 (PARP) and LC3B were determined using a western blot assay in the PC-9 and PC-9/GR cell lines. (B) Endogenous level of EGFR in HCC827, HCC827/GR, H358, and H1650 cells. β-Actin levels were used as loading controls. Column graphs show relative fold change of each protein in gefitinib-treated cells compared with that in control (0; no-gefitinib). Data are mean±standard deviation. *p < 0.05 com pared with control or between indicated group. (Continued to the next page)

Article Snippet: Gefitinib and the antibodies against poly(ADP-ribose) polymerase-1 (PARP), LC3B, integrin-α6, and β-actin were obtained from Cell Signaling Technology, Inc. (Danvers, MA).

Techniques: Incubation, Western Blot, Control, Standard Deviation

Journal: Cancer research and treatment

Article Title: miR-4487 Enhances Gefitinib-Mediated Ubiquitination and Autophagic Degradation of EGFR in Non-Small Cell Lung Cancer Cells by Targeting USP37.

doi: 10.4143/crt.2021.622

Figure Lengend Snippet: Fig. 1. (Continued from the previous page) (C) Reverse transcription polymerase chain reaction was performed to detect the transcriptional activity of EGFR in gefitinib-treated PC-9 cells. (D) PC-9 cells were treated with gefitinib (1 μM) for the indicated time, and total membrane was fractionized and isolated proteins from each fraction were used to detect EGFR levels. Integrin-α6 was used as loading control. PM, plasma membrane; TM, total membrane. (E) Cells were pretreated with gefitinib (1 μM) before the experiments, and PC-9 cells loaded with Fura-2AM were used to determine intracellular Ca2+ ([Ca2+]i) response to epidermal growth factor (EGF; 200 ng/mL) in the absence of extracellular Ca2+. Each trace represents [Ca2+]i mobilization in a single cell.

Article Snippet: Gefitinib and the antibodies against poly(ADP-ribose) polymerase-1 (PARP), LC3B, integrin-α6, and β-actin were obtained from Cell Signaling Technology, Inc. (Danvers, MA).

Techniques: Reverse Transcription, Polymerase Chain Reaction, Activity Assay, Membrane, Isolation, Control, Clinical Proteomics

Journal: Cancer research and treatment

Article Title: miR-4487 Enhances Gefitinib-Mediated Ubiquitination and Autophagic Degradation of EGFR in Non-Small Cell Lung Cancer Cells by Targeting USP37.

doi: 10.4143/crt.2021.622

Figure Lengend Snippet: Fig. 4. miR-4487 directly targets ubiquitin-specific peptidase 37 (USP37) and promotes epidermal growth factor receptor (EGFR) degrada tion and ubiquitination. (A) Differential expression of endogenous miR-4487 was evaluated in BEAS-2B and non–small cell lung cancer (NSCLC) cells using TaqMan primers. Statistical data were presented as fold change of miR-4487 levels in the tested cell lines compared to that in BEAS-2B (control) cells. Data are represented as mean±standard deviation (SD). *p < 0.05 compared with BEAS-2B cells. (B) Endogenous level of miR-4487 was measured in gefitinib (1 μM)-treated PC-9 cells using TaqMan primers. Statistical data were presented as fold change in miR-4487 levels in gefitinib-treated cells compared to that in control (0; no-gefitinib). Data are represented as mean±SD. *p < 0.05 compared with non–gefitinib-treated cells. (C) PC-9 and PC-9/GR cells were transfected with miR-4487 mimic and inhibitor. Total RNA was used for quantitative polymerase chain reaction analysis (left panel). The wild-type (WT) and mutant sequences of USP37 3′- untranslated region (3′-UTR) are listed for comparison. HEK293 cells were co-transfected with either WT or mutated 3′-UTR plasmid and with miR-4487 mimic. Luciferase activity was measured at 48 hours after transfection. Firefly luciferase activity was normalized to Renilla luciferase activity and presented as relative to control (Luc-WT USP37 3′-UTR w/o miR-4487). Data are represented as mean±SD. *p < 0.05 compared to control. I, miR-4487 inhibitor; M, miR-4487 mimic; NC, negative control. (D) miR-4487 mimic and inhibitor was transfected into NSCLC cell lines and endogenous USP37 expression was determined using the western blot assay. (Continued to the next page)

Article Snippet: Gefitinib and the antibodies against poly(ADP-ribose) polymerase-1 (PARP), LC3B, integrin-α6, and β-actin were obtained from Cell Signaling Technology, Inc. (Danvers, MA).

Techniques: Ubiquitin Proteomics, Quantitative Proteomics, Control, Standard Deviation, Transfection, Real-time Polymerase Chain Reaction, Mutagenesis, Comparison, Plasmid Preparation, Luciferase, Activity Assay, Negative Control, Expressing, Western Blot

Journal: Cancer research and treatment

Article Title: miR-4487 Enhances Gefitinib-Mediated Ubiquitination and Autophagic Degradation of EGFR in Non-Small Cell Lung Cancer Cells by Targeting USP37.

doi: 10.4143/crt.2021.622

Figure Lengend Snippet: Fig. 4. (Continued from the previous page) (E) After transfection with NC, M, and I, PC-9 cells were exposed to gefitinib (1 μM) for 24 hours. The endogenous levels of EGFR and LC3B cleavage were analyzed using a western blot assay. (F) miR-4487 mimic was transfected into PC-9 cells and then exposed to gefitinib (1 μM) for 24 hours. Whole-cell ubiquitination was assessed using the UBI-QAPTURE-Q Kit.

Article Snippet: Gefitinib and the antibodies against poly(ADP-ribose) polymerase-1 (PARP), LC3B, integrin-α6, and β-actin were obtained from Cell Signaling Technology, Inc. (Danvers, MA).

Techniques: Transfection, Western Blot, Ubiquitin Proteomics

Journal: Nature Communications

Article Title: LRIG1 is a gatekeeper to exit from quiescence in adult neural stem cells

doi: 10.1038/s41467-021-22813-w

Figure Lengend Snippet: a Schematics of Lrig1 gene disruption. b PCR confirming CRISPR-Cas9 (control and parental line). c Flow cytometry plots with the transfected NSCs. d ICC for LRIG1(red) and nuclear counterstaining with DAPI (blue) in sorted populations after transfection. e EdU quantification of the WT and Lrig1 KO NSCs in the different conditions (EGF/FGF2, BMP, and BMP/FGF2) ( n = 3). f Quantification of the single-cell colony formation in the WT and Lrig1 KO cells in EGF/FGF2 ( n = 3). g Quantification of the number of colonies of deficient and WT NSCs after the treatment with BMP and BMP/FGF2, after to re-exposure to mitogens ( n = 3). h Lrig1 KO cells maintain high levels of pEGFR; WB of EGFR total and EGFR-p in WT and Lrig1 KO cells in the different conditions (EGF/FGF2, BMP and BMP/FGF2). Loading control GAPDH. i Quantification of the EdU positive cells in KO and control cells in the different conditions using Gefitinib ( n = 3 per condition). j Single-cell colony-forming assay of the deficient and control NSCs using Gefitinib ( n = 3 per genotype and condition, 48 single cells plated in each group each time). k Quantification of the colony formation of the WT and Lrig1 KO NSCs in each condition (BMP and BMP/FGF2) using EGFR inhibitor (Gefitinib) ( n = 3 per condition). Scale bar in d = 50um. Data are shown as mean ± SEM of the indicated number of the experiments ( n ) (∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001). Source data are provided as Source Data File.

Article Snippet: BMP-4 (10 ng/ml, Peprotech, AF-120-05ET-100), FGF2 (10 ng/ml, Peprotech, # 100-18b, EGF (10 ng/ml, Peprotech, #315-09), Gefitinib (LKT laboratories, G1721).

Techniques: Disruption, CRISPR, Control, Flow Cytometry, Transfection

Journal: Oncogene

Article Title: A loop of cancer-stroma-cancer interaction promotes peritoneal metastasis of ovarian cancer via TNFα-TGFα-EGFR

doi: 10.1038/onc.2016.509

Figure Lengend Snippet: Fibroblasts-derived TGF-α promotes colony formation of metastatic ovarian cancer cells in 3D organoid co-culture through activation of EGFR, and TGF-α activates AKT and MAPK signaling in metastatic ovarian cancer cells through EGFR. ( a - c ) Phase-contrast images (100x magnification) of ovarian cancer colonies (SKOV3 or PEO1) were captured on day 8 in 3D organoid cultures, including 3D monoculture of ovarian cancer cells treated with rhTGF-α (100 ng/ml) ( a ); 3D organoid co-culture with WI38 fibroblasts ( b ); and 3D organoid co-culture with NOFs ( c ) (upper panel). Treatment of Gefitinib (1 μ M ) or Erlotinib (1 μ M ) was performed. DMSO was served as vehicle controls. Diameter of cancer colonies in 10 fields was measured (Arbitrary unit) in phase-contrast images. The sizes of cancer colonies in the treatment groups were compared with the DMSO control groups (lower panel). ** P <0.01 and *** P <0.001 (Mann-Whitney U test). The data represents means±s.e.m. from two independent experiments. ( d ) Phosphorylation of EGFR, AKT, ERK1/2, STAT1, STAT3 and YAP were examined in ovarian cancer cells (SKOV3 and PEO1) after treatment of rhTGF-α in a time-dependent manner by Western blotting. β-actin was served as loading controls. ( e ) Phosphorylation of EGFR, AKT, ERK1/2 were examined in ovarian cancer cells (SKOV3 and PEO1) after treatments with 100 ng/ml rhTGF-α±1 μ M TKIs (AG1478, Gefitinib or Erlotinib) by Western blotting. DMSO was served as controls. SKOV3 and PEO1 were pre-treated with TKIs for 2 h before stimulating with rhTGF-α for 15 min.

Article Snippet: To investigate the effect of tyrosine kinase inhibitors, the 3D organoid monocultures or 3D organoid co-cultures were treated with 1 μ M of AG1478 (Sigma-Aldrich, Saint Louis, MO, USA), Gefitinib (Toronto Research Chemicals, Canada), or Erlotinib (Toronto Research Chemicals).

Techniques: Derivative Assay, Co-Culture Assay, Activation Assay, MANN-WHITNEY, Western Blot

Journal: Oncogene

Article Title: A loop of cancer-stroma-cancer interaction promotes peritoneal metastasis of ovarian cancer via TNFα-TGFα-EGFR

doi: 10.1038/onc.2016.509

Figure Lengend Snippet: Stromal fibroblasts promote peritoneal metastasis of ovarian cancer in vivo through activation of EGFR, and a TNFα-TGFα-EGFR interacting loop presents in the microenvironment of omental metastases of ovarian cancer. ( a ) Schematic diagram showing the use of orthotopic ovarian cancer xenograft model to evaluate the effect of stromal fibroblasts on peritoneal metastasis of ovarian cancer in vivo , and the effect of EGFR activation on fibroblast-induced peritoneal metastasis of ovarian cancer in vivo . ( b ) Macroscopical morphology of metastatic tumor nodules in orthotopic xenografts was shown (green arrow and green stroke). The weight and number of dissected metastatic tumor nodules were quantified (right). The weight and number of metastatic tumor nodules were compared between the saline-treated mice injected SKOV3+WI38 ( n =5) and the saline-treated mice injected with SKOV3 alone ( n =5), and also between the Gefitinib-treated mice injected with SKOV3+WI38 ( n =5) and the saline-treated mice injected with SKOV3+WI38 ( n =5). * P <0.05 (Student's t test). ( c ) Immunohistochemical staining for TNF-α, TGF-α, EGFR, CD44 and α-SMA in serial sections of metastatic tumor nodules collected from the saline-treated mice injected with SKOV3 alone or with SKOV3+WI38. Fibroblasts were identified by positive signal of α-SMA and CD44 staining. Image was captured at 200x magnification. (Red arrow: cancer cells and black arrow: fibroblasts) ( d ) Immunohistochemical staining for TNF-α, TGF-α, EGFR, and α-SMA in serial sections of omental metastases from patients with advanced ovarian cancer. Image was captured at 200x magnification. (T: ovarian tumor cells and F: omental stromal fibroblasts).

Article Snippet: To investigate the effect of tyrosine kinase inhibitors, the 3D organoid monocultures or 3D organoid co-cultures were treated with 1 μ M of AG1478 (Sigma-Aldrich, Saint Louis, MO, USA), Gefitinib (Toronto Research Chemicals, Canada), or Erlotinib (Toronto Research Chemicals).

Techniques: In Vivo, Activation Assay, Injection, Immunohistochemical staining, Staining