Journal: Nature Communications

Article Title: Prolonging lung cancer response to EGFR inhibition by targeting the selective advantage of resistant cells

doi: 10.1038/s41467-025-61788-w

Figure Lengend Snippet: A Cell viability assay of PC9 cells treated for 5 d with pemetrexed (Pem) and gefitinib (1 µM; Gef). The mean ± SEM of n = 6 is shown (representative of three independent experiments). B PC9 cells containing a pool of EGFR-T790M barcoded cells were treated with gefitinib (1 µM) or pemetrexed (50 nM) for 6 d. The EGFR-T790M mean ± SEM is shown ( n = 4; representative of three independent experiments). C PC9, HCC4006 or HCC827 NSCLC cells containing EGFR-T790M barcoded subpopulations were treated with gefitinib (1 µM) or sorafenib (5 µM; Soraf) for 5 d. The EGFR-T790M mean ± SEM is shown ( n = 4; representative of three independent experiments). D PC9, H1975, and YU-1150 cells containing EGFR-C797S-barcoded subpopulations were treated with osimertinib (0.1 µM; Osim) alone or with sorafenib (5 µM) for 10 d. The EGFR-C797S mean ± SEM is shown ( n = 4; representative of three independent experiments). E PC9 and H1975 cells described in ( E ) were treated with osimertinib (1 µM or 0.1 µM, respectively) alone or with sorafenib (5 µM) for 20 d or 15 d. The cells were then fixed and stained ( n = 3; representative of two and three independent experiments). EGFR-scores of responders ( R ) and non-responders (NR) from a cohort of renal-cell carcinoma ( F ) and thyroid cancer patients ( G ) treated with sorafenib. H H358 cells containing a KRAS-G12D-barcoded subpopulation were treated with sotorasib (10 nM; Sotor) alone or with sorafenib (5 μM) for 12 d. The KRAS-G12D mean ± SEM is shown ( n = 4, representative of three independent experiments). p = 0.4857. I Ceritinib-resistant H3122 cells labeled with VIRHD lentivirus were mixed with parental cells (1:50) and treated with ceritinib (50 nM; Cerit) alone or with sorafenib (5 µM) for 7 d. VIRHD-cells mean ± SEM is shown ( n = 5, representative of three independent experiments). J EGFR-C797S CRISPR-barcoded PC9 cells were treated with osimertinib (0.1 µM) alone or with sorafenib (5 µM), sunitinib (1 µM; Sun), regorafenib (2 µM; Reg), lenvatinib (1 µM; Lenv), or cabozantinib (5 µM; Cab) for 7 d (left) or 10 d (right). EGFR-C797S mean ± SEM is shown ( n = 4–5; representative of three independent experiments). K PC9 cells were treated with sorafenib (5 μM) for 72 h, and cell cycle was analyzed by FACS. The mean ± SEM of four independent experiments is shown. L PC9 cells were treated with sorafenib (5 μM) for 72 h, and percentage of apoptotic cells was measured by annexin V staining. The mean ± SEM of three independent experiments is shown. ns not significant. Statistics are: B , C , D , H , I , J matched Mann-Whitney, two-tailed; F , G matched Student t -test, two-tailed; K , L matched two-way ANOVA with the Tukey correction. N refers to biological replicates. Source data are provided as a file.

Article Snippet: Human embryonic kidney 293T cells were obtained from ATCC; PC9 cells (NSCLC, EGFR-Ex19Del) were obtained from ECACC (distributed by Sigma-Aldrich); HCC4006 (NSCLC, EGFR-Ex19Del) and H1975 cells (NSCLC, EGFR-L858R/T790M) were obtained from ATCC; H3122 cells (NSCLC, EML4-ALK) were obtained from Cytion; HCC827 cells (NSCLC, EGFR-Ex19Del) and H358 (NSCLC, KRAS-G12C) were a gift from Pr.

Techniques: Viability Assay, Staining, Labeling, CRISPR, MANN-WHITNEY, Two Tailed Test

Journal: Nature Communications

Article Title: Prolonging lung cancer response to EGFR inhibition by targeting the selective advantage of resistant cells

doi: 10.1038/s41467-025-61788-w

Figure Lengend Snippet: A PC9 cells containing subpopulations of EGFR-G724S, ERBB2-ex20ins, KRAS-G12D, BRAF-V600E or PIK3CA-E545K cells were treated as indicated with osimertinib (0,1 μM; Osim), sorafenib (5 μM; Soraf) or trametinib (10 nM; Tram) for 7 d to 20 d. The mean ± SEM of the mutant barcodes is shown ( n = 4; representative of three independent experiments). B BRAF-V600E osimertinib-resistant YUX-1024 and YU-1150 cells were mixed with parental cells (1:100) and treated for 7 d with osimertinib (0.1 μM) alone or with sorafenib (5 μM). The BRAF-V600E mean ± SEM is shown ( n = 4; representative of three independent experiments). C PC9 cells containing a EML4-ALK-barcoded subpopulation were treated for 10 d with osimertinib (0.1 µM), sorafenib (5 µM), or crizotinib (500 nM; Criz). The EML4-ALK mean ± SEM is shown ( n = 5; representative of three independent experiments). D PC9 cells overexpressing ERBB2-ex20ins or MET were mixed with parental cells (1:100) and treated with osimertinib (0.1 μM) alone or with sorafenib (5 μM) for 10 d or 15 d. The mean fraction ± SEM of ERBB2-ex20ins- or MET-overexpressing cells is shown ( n = 4; representative of three independent experiments). E Lentiviral-labeled, osimertinib-resistant HCC827-GR6 cells were mixed with parental HCC827 (1:100) and treated for 6 d with osimertinib (0.1 µM) alone or with sorafenib (5 µM). HCC827-GR6 mean fraction ± SEM is shown ( n = 4; representative of three independent experiments). F PC9 cells transduced with empty or inducible-SNAI2 vectors were pre-treated with or without doxycycline (1 µg/ml; Dox) for 7 d, mixed with parental PC9 (1:100) and treated for 7 d with osimertinib (0.1 µM) alone or with sorafenib (5 µM). The mean fraction ± SEM of vector-labeled cells is shown ( n = 4; representative of three independent experiments). LIM1215 cells containing EGFR-G465R ( G ) or KRAS-G12D ( H ) CRISPR-barcodes were treated for 6 d with cetuximab (20 µg; Cetux) or osimertinib (1 µM), alone or with sorafenib (5 μM). The barcode mean fraction ± SEM is shown ( n = 3 for EGFR-G465R; n = 4 for KRAS-G12D; representative of three independent experiments). Statistics calculated by Mann–Whitney test, two-tailed. “ N ” refers to biological replicates. Source data are provided as a file.

Article Snippet: Human embryonic kidney 293T cells were obtained from ATCC; PC9 cells (NSCLC, EGFR-Ex19Del) were obtained from ECACC (distributed by Sigma-Aldrich); HCC4006 (NSCLC, EGFR-Ex19Del) and H1975 cells (NSCLC, EGFR-L858R/T790M) were obtained from ATCC; H3122 cells (NSCLC, EML4-ALK) were obtained from Cytion; HCC827 cells (NSCLC, EGFR-Ex19Del) and H358 (NSCLC, KRAS-G12C) were a gift from Pr.

Techniques: Mutagenesis, Labeling, Transduction, Plasmid Preparation, CRISPR, MANN-WHITNEY, Two Tailed Test

Journal: Scientific Reports

Article Title: GFP-complementation assay to detect functional CPP and protein delivery into living cells

doi: 10.1038/srep18329

Figure Lengend Snippet: ( 3a,b) SEE components are validated by co-transfection of CHO-K1 cells with hGFP1-10 and various Cargo_S11 fusion constructs. All S11 fusions complemented hGFP1-10, measured by increasing % fluorescent cells ( 3a ) or increasing MFI ( 3b ) in the viable single-cell population. In all experiments the “S11 only” control is a fusion of the [GSSG]x2 linker and S11 sequence, and does not generate a detectable GFP signal, nor do empty vector and “GFP1-10 only” controls (% GFP-positive cells <0.65 or MFI <25). Error bars represent standard error of the mean between technical replicates; data is representative of three independent experiments. ( 3c) Fluorescence microscopy detects GFP complementation (FITC channel) in HCC827/hGFP1-10 cell populations transfected with complementing β-ACTIN_S11 or TRX_S11 constructs, compared to empty vector control. Cells are also counter-stained for endogenous β-Actin (TRITC) and nuclei (DAPI) before visualizing. Bar scale is 50 μm.

Article Snippet: CHO-K1 and HCC827 monoclonal cell lines stably expressing hGFP1-10 were made through limiting dilution by Genecopoeia (USA), and were maintained in Zeocin-supplemented parental cell line medium with 200 ug/ml and 250 ug/ml Zeocin, respectively.

Techniques: Cotransfection, Construct, Control, Sequencing, Plasmid Preparation, Fluorescence, Microscopy, Transfection, Staining

Journal: Scientific Reports

Article Title: GFP-complementation assay to detect functional CPP and protein delivery into living cells

doi: 10.1038/srep18329

Figure Lengend Snippet: ( 4a,b) CHO-K1 cells are co-transfected with hGFP1-10 and various Cargo-linker combinations. All_S11 fusions complemented hGFP1-10 to various extents, measured by increasing % fluorescent cells ( 4a ) or increasing MFI ( 4b ) in the viable single-cell population. In all experiments the “S11 only” control is a fusion of the [GSSG]x2 linker and S11 sequence, and does not generate a detectable GFP signal, nor do empty vector and “GFP1-10 only” controls (% GFP-positive cells <0.65 or MFI <25). Error bars represent standard error of the mean between technical replicates; data is representative of three independent experiments. ( 4c–f) Transfection experiments validating CPP components were repeated in a monoclonal stable cell line: HCC827 cells expressing hGFP1-10. HCC827/hGFP1-10 cells are transfected with different Cargo_S11 fusion constructs ( 4c,d ) or various Cargo-linker combinations ( 4e,f ). All Cargo_S11 proteins and various Cargo-linker combinations complemented hGFP1-10, as measured by increasing % GFP-positive cells ( 4c,e ) or increasing MFI ( 4d,f ) in the viable single-cell population and compared to background (no GFP1-10 and/or empty vector). In all experiments the “S11 only” control is expressed [GSSG]x2 linker_S11 fusion and does not generate a detectable increase in GFP signal, nor do empty vector and “GFP1-10 only” controls (% GFP-positive cells <5%, or MFI <110). Error bars represent standard error of the mean between technical replicates; data is representative of three independent experiments.

Article Snippet: CHO-K1 and HCC827 monoclonal cell lines stably expressing hGFP1-10 were made through limiting dilution by Genecopoeia (USA), and were maintained in Zeocin-supplemented parental cell line medium with 200 ug/ml and 250 ug/ml Zeocin, respectively.

Techniques: Transfection, Control, Sequencing, Plasmid Preparation, Stable Transfection, Expressing, Construct

Journal: Scientific Reports

Article Title: GFP-complementation assay to detect functional CPP and protein delivery into living cells

doi: 10.1038/srep18329

Figure Lengend Snippet: CPP_TRX_S11 proteins were added to CHO-K1 ( 5a,b ) and HCC827 ( 5c,d ) monoclonal cell lines stably expressing hGFP1-10. Protein titration shows dose-dependent uptake for TAT, Penetratin (PEN), Penetratin-Arginine (PenArg) and R9 fusions read as GFP complementation measured by both % fluorescent cells ( 5a,c ) and fold change in MFI ( 5b,d ). The seven other canonical CPPs showed only minimal GFP complementation signal at the highest doses. Control TRX_S11 fusion proteins “No CPP” and “PYC35” (a peptide with no CPP activity) show negligible effect on GFP complementation signal over cell-line background (“HisMBP”). Error bars represent standard error of the mean between technical replicates; data is representative of two independent experiments per cell line. (*R9 and PenArg fusions were not sufficiently concentrated for testing at 40 μM.)

Article Snippet: CHO-K1 and HCC827 monoclonal cell lines stably expressing hGFP1-10 were made through limiting dilution by Genecopoeia (USA), and were maintained in Zeocin-supplemented parental cell line medium with 200 ug/ml and 250 ug/ml Zeocin, respectively.

Techniques: Stable Transfection, Expressing, Titration, Control, Activity Assay

Journal: Scientific Reports

Article Title: GFP-complementation assay to detect functional CPP and protein delivery into living cells

doi: 10.1038/srep18329

Figure Lengend Snippet: Recombinant proteins are added to CHO-K1 and HCC827 cells transiently transfected 24-hours prior with hGFP1-10 and left overnight to recover and express the protein. GFP complementation was measured as both % fluorescent cells ( 8a ) and fold change in MFI ( 8b ). A dose-dependent fluorescent signal is detected for the TAT fusion protein compared to controls illustrating CPP-dependent internalization in transfected cells. “No CPP” control is the TRX_S11 protein with no CPP moiety added. “PYC35” control is an unrelated peptide (known to have no CPP activity) instead of a CPP moiety expressed as part of the fusion protein. Data is representative of more than 10 independent experiments. Error bars represent standard error of the mean between technical replicates.

Article Snippet: CHO-K1 and HCC827 monoclonal cell lines stably expressing hGFP1-10 were made through limiting dilution by Genecopoeia (USA), and were maintained in Zeocin-supplemented parental cell line medium with 200 ug/ml and 250 ug/ml Zeocin, respectively.

Techniques: Recombinant, Transfection, Control, Activity Assay

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Quercetin Inhibits the Proliferation and Metastasis of Human Non-Small Cell Lung Cancer Cell Line: The Key Role of Src-Mediated Fibroblast Growth Factor-Inducible 14 (Fn14)/Nuclear Factor kappa B (NF-κB) pathway

doi: 10.12659/MSM.920537

Figure Lengend Snippet: Administration of Que suppressed the proliferation potential of HCC827 cells. For MTT assays, cells were incubated with Que of 25 μM (Que L), 50 μM (Que M), and 100 μM (Que H) for 96 hours. Every 24 hours, cells were subjected to the testing. For colony formation assays, cells were cultured in medium containing 100 μM Que for 2 weeks. ( A ) Detection results of MTT assays. ( B ) Detection results of colony formation assays. * P <0.05 versus the Control group.

Article Snippet: Human NSCLC cell lines HCC827 (7-1150) and NCI-H1650 (7-1031) were obtained from Chi Scientific (MA, USA).

Techniques: Incubation, Cell Culture, Control

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Quercetin Inhibits the Proliferation and Metastasis of Human Non-Small Cell Lung Cancer Cell Line: The Key Role of Src-Mediated Fibroblast Growth Factor-Inducible 14 (Fn14)/Nuclear Factor kappa B (NF-κB) pathway

doi: 10.12659/MSM.920537

Figure Lengend Snippet: Administration of Que inhibited the invasion and migration potentials, and the signaling transduction of Src/Fn14/NF-κB pathway in HCC827 cells. For scratch assays, cells were incubated with Que of 100 μM (Que H) for 48 hours. For Transwell assays, cells were incubated with Que of 100 μM (Que H) for 24 hours. For western blotting assays, cells were incubated with Que of 100 μM (Que H) for 24 hours. ( A ) Detection results of scratch assays. ( B ) Detection results of Transwell assays. ( C ) Detection results of western blotting assays. * P <0.05 versus the Control group.

Article Snippet: Human NSCLC cell lines HCC827 (7-1150) and NCI-H1650 (7-1031) were obtained from Chi Scientific (MA, USA).

Techniques: Migration, Transduction, Incubation, Western Blot, Control

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Quercetin Inhibits the Proliferation and Metastasis of Human Non-Small Cell Lung Cancer Cell Line: The Key Role of Src-Mediated Fibroblast Growth Factor-Inducible 14 (Fn14)/Nuclear Factor kappa B (NF-κB) pathway

doi: 10.12659/MSM.920537

Figure Lengend Snippet: Overexpression of Src blocked the anti-NSCLC function of Que in HCC827 cells. Cells transfected with negative control vector (NC) or Src expression vector (Src) were treated with Que of 100 μM (Que H) and subjected to MTT assay for 96 hours (cells were collected every 24 hours), and colony formation was assay at 2 weeks, Transwell assay at 24 hours, and scratch assay at 48 hours. ( A ) Detection results of Src level. ( B ) Detection results of MTT assays. ( C ) Detection results of colony formation assay. ( D ) Detection results of Transwell assay. ( E ) Detection results of scratch assay. # P <0.05 versus Que H group.

Article Snippet: Human NSCLC cell lines HCC827 (7-1150) and NCI-H1650 (7-1031) were obtained from Chi Scientific (MA, USA).

Techniques: Over Expression, Transfection, Negative Control, Plasmid Preparation, Expressing, MTT Assay, Transwell Assay, Wound Healing Assay, Colony Assay

Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

Article Title: Quercetin Inhibits the Proliferation and Metastasis of Human Non-Small Cell Lung Cancer Cell Line: The Key Role of Src-Mediated Fibroblast Growth Factor-Inducible 14 (Fn14)/Nuclear Factor kappa B (NF-κB) pathway

doi: 10.12659/MSM.920537

Figure Lengend Snippet: Que inhibited the growth and metastasis potential of NSCLC in vivo in by inhibiting Src signaling. Mice were injected with different HCC827 cells and administrated with Que of 100 mg/kg body weight for 3 weeks. ( A ) Detection results of tumor volume. ( B ) Detection results of hematoxylin and eosin detection of tumor tissue. ( C ) Detection results of western blotting detection of E-cadherin and N-cadherin. * P <0.05 versus Que H+Src group. Scale bar, 100 μm.

Article Snippet: Human NSCLC cell lines HCC827 (7-1150) and NCI-H1650 (7-1031) were obtained from Chi Scientific (MA, USA).

Techniques: In Vivo, Injection, Western Blot

Journal: American Journal of Translational Research

Article Title: Fisetin, a dietary phytochemical, overcomes Erlotinib-resistance of lung adenocarcinoma cells through inhibition of MAPK and AKT pathways

doi:

Figure Lengend Snippet: A: The chemical structure of fisetin. B: Characterization of HCC827 and HCC827-ER cells. Western blot analysis showed p-MAPK, p-Akt and Survivin were up-regulated in HCC827-ER compared to HCC827cells (P<0.05). C: Western blot analysis showed p-AXL and Snail were up-regulated while E-cadherin was down-regulated in HCC827-ER compared to HCC827cells. D: Effects of Fisetin on the viability of HCC827-ER cells. Cells were treated with Fisetin (0-120 μM) for 24 and 48 h. Viability was determined by MTT assay. The data is presented as mean ± SD of three separate experiments. *P<0.05, vs control group (0 μM).

Article Snippet: An acquired Erlotinib-resistant subclone cell line of HCC827 was established in continuous culture with gradually increasing Erlotinib (Genentech, San Francisco, CA) [ 26 ].

Techniques: Western Blot, MTT Assay, Control

Journal: American Journal of Translational Research

Article Title: Fisetin, a dietary phytochemical, overcomes Erlotinib-resistance of lung adenocarcinoma cells through inhibition of MAPK and AKT pathways

doi:

Figure Lengend Snippet: Cells were divided into four subgroups and treated with 40 μM Fisetin, 15 μM Erlotinib, 40 μM Fisetin + 15 μM Erlotinib, and DMEM as a control for 24 h, respectively. Then, cell viability and apoptosis were tested. Cell viability (A) and apoptosis (B) in HCC827-ER assessed by MTT and apoptosis assay. (*P<0.05 vs Control. †P<0.05 vs Control) Expression of the MAPK, AKT, Survivin and apoptosis pathway-related proteins (C) and AXL, Snail and E-cadherin proteins (D) assessed by immunoblotting.

Article Snippet: An acquired Erlotinib-resistant subclone cell line of HCC827 was established in continuous culture with gradually increasing Erlotinib (Genentech, San Francisco, CA) [ 26 ].

Techniques: Control, Apoptosis Assay, Expressing, Western Blot

Journal: American Journal of Translational Research

Article Title: Fisetin, a dietary phytochemical, overcomes Erlotinib-resistance of lung adenocarcinoma cells through inhibition of MAPK and AKT pathways

doi:

Figure Lengend Snippet: Effects of Fisetin and Erlotinib on the growth inhibition of HCC827-ER. Cell growth inhibition curves of HCC827-ER cells treated with Fisetin, Erlotinib or the combination of Fisetin and Erlotinib for 24 h (A) or 48 h (B). Analysis of combined effects of Fisetin and Erlotinib on HCC827-ER cells for 24 h (C) or 48 h (D) treatment. CI values were determined according to the median-effect method by Chou and Talalay, with less than 1 at all given concentrations indicating highly synergistic effects. The molar ratios of Fisetin and Erlotinib were fixed at 1:1 with increasing doses (from 10 μM to 120 μM), respectively. Data were presented as means ± SD, n=3.

Article Snippet: An acquired Erlotinib-resistant subclone cell line of HCC827 was established in continuous culture with gradually increasing Erlotinib (Genentech, San Francisco, CA) [ 26 ].

Techniques: Inhibition

Journal: American Journal of Translational Research

Article Title: Fisetin, a dietary phytochemical, overcomes Erlotinib-resistance of lung adenocarcinoma cells through inhibition of MAPK and AKT pathways

doi:

Figure Lengend Snippet: HCC827-ER cells were divided into eight subgroups as I to VIII, respectively. They were treated with different reagent for 24 h as follows: DMEM (I), Erlotinib (II), U0126 + MK-2206 (III), R428 (IV), Fisetin (V), Erlotinib + U0126 + MK-2206 (VI), Erlotinib + R428 (VII), and Erlotinib + Fisetin (VIII). Cell viability (A) and apoptosis (B) of the cells were assessed by MTT and apoptosis assays, respectively. (*P<0.05 vs I or II; †P<0.05 vs VI or VII).

Article Snippet: An acquired Erlotinib-resistant subclone cell line of HCC827 was established in continuous culture with gradually increasing Erlotinib (Genentech, San Francisco, CA) [ 26 ].

Techniques:

Journal: American Journal of Translational Research

Article Title: Fisetin, a dietary phytochemical, overcomes Erlotinib-resistance of lung adenocarcinoma cells through inhibition of MAPK and AKT pathways

doi:

Figure Lengend Snippet: HCC827-ER cells were divided into four subgroups and treated with 15 μM Erlotinib, 1 μM R428 (a specific AXL inhibitor), 1 μM R428 + 15 μM Erlotinib, and DMEM as a control for 24 h, respectively. Cell viability (A) and apoptosis (B) in HCC827-ER were assessed by MTT and apoptosis assays, respectively. (*P<0.05 vs Control). Expression of the AXL, MAPK, AKT, Survivin and apoptosis pathway-related proteins were assessed by immunoblotting.

Article Snippet: An acquired Erlotinib-resistant subclone cell line of HCC827 was established in continuous culture with gradually increasing Erlotinib (Genentech, San Francisco, CA) [ 26 ].

Techniques: Control, Expressing, Western Blot

Journal: The Journal of Biological Chemistry

Article Title: Rho family small GTPase Rif regulates Wnt5a-Ror1-Dvl2 signaling and promotes lung adenocarcinoma progression

doi: 10.1016/j.jbc.2023.105248

Figure Lengend Snippet: Ror1 and Rif are colocalized at filopodia in LUAD cells. A, Western blot analysis of Ror1, Ror2, and Rif in LUAD cell lines, showing expression of both Ror1 and Rif were detectable in all LUAD cell lines examined. Blots are representative of two independent experiments. B, coimmunoprecipitation assay, showing association between Rif and Ror1 at endogenous protein levels in the indicated LUAD cells. Whole-cell lysates were subjected to immunoprecipitation (IP) with anti-Ror1 antibody or control immunoglobulin G (isotype matched), followed by Western blotting. Blots are representative of six (PC9), three (HCC827), and two (A549) independent experiments. C and D, representative xz-images of PC9 cells expressing YFP-Rif and Ror1-mCh on a 2D surface ( C ) or Matrigel ( D ), showing colocalization of these proteins at filopodia. Phalloidin staining in ( D ) shows highly accumulated F-actin on the front side containing filopodia. In ( D ), the xy images ( lower panels ) sectioned along the white arrow in xz confocal image ( upper panels ) are shown. Images are representative of at least three independent experiments. The scale bars represent 10 μm. Magnified images of boxed regions are shown on the right . The scale bars in magnified images represent 2 μm ( C ) and 5 μm ( D ). LUAD, lung adenocarcinoma; Rif, Rho in filopodia.

Article Snippet: Human LUAD cell lines PC9, A110L, H1993, and H1975 were provided by Y. Maniwa (Kobe University), and HCC827 (which express luciferase stably) and A549 cells were purchased from JCRB cell bank.

Techniques: Western Blot, Expressing, Co-Immunoprecipitation Assay, Immunoprecipitation, Control, Staining

Journal: Molecular cancer therapeutics

Article Title: TAK-701, a humanized monoclonal antibody to HGF, reverses gefitinib resistance induced by tumor-derived HGF in non-small cell lung cancer with an EGFR mutation

doi: 10.1158/1535-7163.MCT-10-0481

Figure Lengend Snippet: Characterization of HCC827 isogenic cell lines. A, HCC827 isogenic cell lines (HCC827, HCC827-Mock, HCC827-HGF1 and -HGF2, and HCC827 GR5) were cultured overnight in medium containing 10% serum and then incubated for 24 h in serum-free medium, after which the culture supernatants were collected and assayed for HGF with an ELISA. Data are means ± SD from three independent experiments. B, HCC827 isogenic cell lines were cultured in medium containing 10% serum for 72 h in the presence of various concentrations of gefitinib, after which cell viability was assessed as described in Materials and Methods. The number of viable cells is expressed as a percentage of the value for untreated cells. Data are means ± SD from three independent experiments. C, HCC827 isogenic cell lines were incubated for 1 h with or without gefitinib (100 nM) in medium containing 10% serum, after which the cells were lysed and subjected to immunoblot analysis with antibodies to phosphorylated (p-) or total forms of EGFR, MET, AKT, or ERK or with those to β-actin (loading control).

Article Snippet: HCC827 cells were implanted subcutaneously into the right hind leg of 6-week-old female athymic nude mice (BALB/c nu/nu; CLEA Japan, Tokyo, Japan).

Techniques: Cell Culture, Incubation, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Molecular cancer therapeutics

Article Title: TAK-701, a humanized monoclonal antibody to HGF, reverses gefitinib resistance induced by tumor-derived HGF in non-small cell lung cancer with an EGFR mutation

doi: 10.1158/1535-7163.MCT-10-0481

Figure Lengend Snippet: Effects of the combination of gefitinib and either TAK-701 or PHA-665752 on the growth of gefitinib-resistant NSCLC cells. HCC827 GR5 cells (A), HCC827-HGF1 cells (B), or HCC827-HGF2 cells (C) were cultured for 72 h in medium containing 10% serum and various concentrations of gefitinib and either PHA-665752 (500 nM) or TAK-701 (50 μg/mL), after which cell viability was assessed. Data are means ± SD from three independent experiments.

Article Snippet: HCC827 cells were implanted subcutaneously into the right hind leg of 6-week-old female athymic nude mice (BALB/c nu/nu; CLEA Japan, Tokyo, Japan).

Techniques: Cell Culture

Journal: Molecular cancer therapeutics

Article Title: TAK-701, a humanized monoclonal antibody to HGF, reverses gefitinib resistance induced by tumor-derived HGF in non-small cell lung cancer with an EGFR mutation

doi: 10.1158/1535-7163.MCT-10-0481

Figure Lengend Snippet: Effects of the combination of gefitinib and either TAK-701 or PHA-665752 on cell signaling in gefitinib-resistant NSCLC cells. HCC827 cells, HCC827 GR5 cells, or HCC827-HGF2 cells were incubated for 6 h in medium containing 10% serum in the absence or presence of gefitinib (1 μM), PHA-665752 (500 nM), or TAK-701 (50 μg/mL), as indicated. Cell lysates were then prepared and subjected to immunoblot analysis with antibodies to phosphorylated or total forms of EGFR, MET, AKT, or ERK or with those to β-actin.

Article Snippet: HCC827 cells were implanted subcutaneously into the right hind leg of 6-week-old female athymic nude mice (BALB/c nu/nu; CLEA Japan, Tokyo, Japan).

Techniques: Incubation, Western Blot

Journal: Molecular cancer therapeutics

Article Title: TAK-701, a humanized monoclonal antibody to HGF, reverses gefitinib resistance induced by tumor-derived HGF in non-small cell lung cancer with an EGFR mutation

doi: 10.1158/1535-7163.MCT-10-0481

Figure Lengend Snippet: Effects of the combination of TAK-701 and gefitinib on the growth of gefitinib-resistant NSCLC cells in vivo. Nude mice with tumor xenografts established by subcutaneous injection of HCC827-Mock (A) or HCC827-HGF2 (B) cells were treated for 7 weeks with vehicle (control), gefitinib (50 mg/kg), TAK-701 (5 mg/kg), or both drugs, as described in Materials and Methods. Tumor volume was determined at the indicated times after the onset of treatment. Data are means ± SEM from five mice per group.

Article Snippet: HCC827 cells were implanted subcutaneously into the right hind leg of 6-week-old female athymic nude mice (BALB/c nu/nu; CLEA Japan, Tokyo, Japan).

Techniques: In Vivo, Injection