Journal: Cancers

Article Title: Regression of Lung Cancer in Mice by Intranasal Administration of SARS-CoV-2 Spike S1

doi: 10.3390/cancers14225648

Figure Lengend Snippet: Effect of recombinant SARS-CoV-2 spike S1 on the survival of human H1299 and H358 lung cancer cells. H1299 ( A , C ) and H358 ( B , D ) cells were treated with spike S1 protein for 24 h under serum-free condition followed by monitoring cell death by LDH release ( A , B ) and MTT ( C , D ). Results are mean + SD of three different experiments. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Same procedure was utilized for two other cell lines purchased from ATCC: H1299 (human NSCLC, p53 negative; Catalog# CRL-5803) H358 (human NSCLC, KRAS mutant; Catalog# CRL-5807) These cells were cultured in RPMI-1640 containing 10% FBS, 100 U/mL penicillin, and 100 µg/mL streptomycin.

Techniques: Recombinant

Journal: Journal of Translational Medicine

Article Title: IκBα kinase inhibitor BAY 11-7082 promotes anti-tumor effect in RAS-driven cancers

doi: 10.1186/s12967-024-05384-4

Figure Lengend Snippet: BAY 11-7082 treatment alters the expression of multiple genes in NRAS, KRAS, and HRAS mutant cancer cells. (A) Heatmaps showing the top 50 upregulated or downregulated genes in SKMEL-103, AsPC1, and RH-36 cells upon treatment with BAY 11-7082 (5 µM) for 48 h as compared with DMSO-treated cells. (B) Volcano plot showing top 15 genes upregulated or downregulated after 48-h treatment with BAY 11-7082 (5 µM) in RAS mutant cells

Article Snippet: RAS mutant cancer cell lines (NRAS: M245, SKMEL-103, SKMEL-2; KRAS: PANC1, AsPC1, SU.86.86) were purchased from American Type Culture Collection (ATCC) as listed in Supplementary Tables and maintained in a humidified atmosphere of 5% CO2 at 37 °C in Dulbecco’s modified Eagle medium (Life Technologies, Carlsbad, CA, USA) or Roswell Park Memorial Institute-1640 Medium (Life Technologies), each supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin (both from Life Technologies).

Techniques: Expressing, Mutagenesis

Journal: Journal of Translational Medicine

Article Title: IκBα kinase inhibitor BAY 11-7082 promotes anti-tumor effect in RAS-driven cancers

doi: 10.1186/s12967-024-05384-4

Figure Lengend Snippet: Model showing the new IκBα kinase Inhibitor BAY 11-7082 for Treating RAS-driven cancers. A model depicting various cancer types driven by oncogenic NRAS, KRAS, and HRAS mutations overexpress IκBα kinase. IκBα kinase can be effectively targeted by small molecule inhibitor BAY 11-7082, leading to downregulation of several prooncogenic signaling pathways, including PI3K-AKT signaling cascade and upregulation of apoptosis that contributes to tumor growth inhibition observed in RAS-driven cancers

Article Snippet: RAS mutant cancer cell lines (NRAS: M245, SKMEL-103, SKMEL-2; KRAS: PANC1, AsPC1, SU.86.86) were purchased from American Type Culture Collection (ATCC) as listed in Supplementary Tables and maintained in a humidified atmosphere of 5% CO2 at 37 °C in Dulbecco’s modified Eagle medium (Life Technologies, Carlsbad, CA, USA) or Roswell Park Memorial Institute-1640 Medium (Life Technologies), each supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin (both from Life Technologies).

Techniques: Protein-Protein interactions, Inhibition

Journal: Communications Biology

Article Title: Secreted midbody remnants are a class of extracellular vesicles molecularly distinct from exosomes and microparticles

doi: 10.1038/s42003-021-01882-z

Figure Lengend Snippet: a Venn diagram of proteins identified in SW620 cell-derived sMB-R (sMV-HD), sMV-LD and Exos. b Heatmap illustration of proteins identified in sMB-R (sMV-HD), sMV-LD and Exos. Proteins present in higher abundance in sMB-R (red) as compared to sMV-LD and Exos include conserved cytokinetic proteins as well as additional cytokinetic proteins. *Proteins uniquely identified in sMB-Rs. **Proteins enriched (fold change >2) in sMB-R compared to sMV-LD and Exos. c STRING-based protein-protein interaction network analysis of 928 enriched proteins in sMB-Rs (sMV-HD) compared to sMV-LD and Exos. The interactions were “evidence”-based, with “experiments” as active interaction source and interaction threshold set at 0.900 (highest confidence). Disconnected nodes in the network are hidden. Proteins identified under biological processes or molecular processes (Gene Ontology) are indicated. Centralspindlin complex components (RACGAP1 and KIF23/MKLP1) are also indicated. d EnrichmentMap of Reactome pathways enriched in 456 proteins commonly identified in SW620 cell-derived sMB-R proteome (2300 proteins) with the proteome of MB-Rs shed by Hela cells reported recently by Peterman et al. 2019 . e Mass spectrometry-based identification of KRAS peptides (UniProtKB ID RASK_HUMAN) in sMB-Rs. Two peptides (TEYKLVVVGAGGVGK and LVVVGAGGVGK) spanning Gly-12/ Val-12 substitution in KRAS protein. Peptide spectral profiles are displayed on the right. f Immunofluorescence microscopy of SW620 cells using anti-MKLP1 and anti-KRAS G12V antibodies. Nuclei (blue) were stained with Hoechst stain. White arrows indicate the position of MB and MB-Rs. Inset represents higher magnification. Scale bar, 10 µm. g Western blot analysis of exosomes, crude 10,000 x g sMVs, and isopycnic (iodixanol-density) gradient centrifugation fractions of sMV-LD and -HD/sMB-Rs using anti-KRAS G12V antibody.

Article Snippet: Cells were then incubated with primary antibodies (1:100) (mouse anti-MKLP1 (Santa Cruz Biotechnology), mouse anti-RACGAP1 (Santa Cruz), rabbit anti-KRAS G12V mutant specific (Cell Signalling), rabbit anti-RAB7 (Abcam) and rabbit anti-β-tubulin (Cell Signalling) in blocking solution for 1 h at room temperature.

Techniques: Derivative Assay, Mass Spectrometry, Immunofluorescence, Microscopy, Staining, Western Blot, Gradient Centrifugation

Journal: Communications Biology

Article Title: Secreted midbody remnants are a class of extracellular vesicles molecularly distinct from exosomes and microparticles

doi: 10.1038/s42003-021-01882-z

Figure Lengend Snippet: a Uptake of sMB-Rs by fibroblasts. Fluorescence microscopy analysis of NIH3T3 fibroblasts incubated with/without SW620 cell-derived sMB-Rs or Exos (50 µg ml −1 ) for 2 h using anti-MKLP1 and anit-RACGAP1 antibodies. b Uptake and accumulation of sMB-Rs in NIH3T3 fibroblasts was quantified by counting MKLP1 + puncta per cell; data represented as mean ± s.e.m. Nuclei (blue) were stained with Hoechst. Scale bar, 10 µm. c Internalisation of sMB-Rs by fibroblasts. Confocal microscopy of NIH3T3 fibroblasts incubated with sMB-Rs using anti-MKLP1 (in green) and anti-RAB7 (in red) antibodies. Confocal microscopy analysis along Z-axis (inset) reveal internalisation of sMB-Rs following uptake. Scale bar, 10 µm. d Intercellular transfer of sMB-R KRAS G12V into NIH3T3 cells. Fluorescence microscopy of NIH3T3 fibroblasts incubated with SW620 cell-derived sMB-Rs (5 µg) for 2 h using anti-KRAS G12V antibodies. Nuclei were stained with Hoechst stain (blue). Right panel represents fluorescence signals from left panel overlaid onto bright-field images. Inset represents enlarged image. Scale bar, 10 µm.

Article Snippet: Cells were then incubated with primary antibodies (1:100) (mouse anti-MKLP1 (Santa Cruz Biotechnology), mouse anti-RACGAP1 (Santa Cruz), rabbit anti-KRAS G12V mutant specific (Cell Signalling), rabbit anti-RAB7 (Abcam) and rabbit anti-β-tubulin (Cell Signalling) in blocking solution for 1 h at room temperature.

Techniques: Fluorescence, Microscopy, Incubation, Derivative Assay, Staining, Confocal Microscopy

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: The expression of GSTP1 and p53 in PC specimens. (A) Low and high GSTP1 expression in two normal pancreas samples. (B) Low and high GSTP1 expression in two PC samples. (C) Low and high p53 expression in two PC samples. (D) High GSTP1 and mtp53 expression in one PC sample. (E) Low GSTP1 and mtp53 expression in one PC sample. (F) High GSTP1 and wtp53 expression in one serial section of PC tissue. (G) Kaplan–Meier analysis of high and low expression of GSTP1 in PC patients. (H) Kaplan–Meier analysis of Mtp53 and wtp53 expression in PC patients. (I) Kaplan–Meier analysis of high and low expression of GSTP1 in wtp53 PC patients.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: Expressing

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: The relationship between GSTP1 and p53 in vitro. (A) GSTP1 and p53 protein expression in six PC cell lines. (B) GSTP1 mRNA expression in six PC cell lines. (C) GSTP1 and p53 expression in GSTP1‐ or p53‐silenced mtp53 BxPC‐3 cells. (D) GSTP1 and p53 expression in GSTP1‐ or p53‐silenced mtp53 PANC‐1 cells. (E) GSTP1 and p53 expression in GSTP1‐ or p53‐silenced wtp53 Capan‐2 cells. (F) GSTP1 and p53 expression in the GFP, GSTP1‐GFP, GSTP1‐GFP plus si‐p53, and si‐p53 groups of SW1990 cells with wtp53. 1: sictrl group; 2: si1‐GSTP1 group; 3: si2‐GSTP1 group; 4: si‐p53 group; 5: GFP group; 6: GSTP1‐GFP group; 7: GSTP1‐GFP + si‐p53 group; 8: Si‐p53 group. The experiments were performed in triplicate. Error bars ± SDs. * p < 0.05 and ** p < 0.01 compared with the control.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: In Vitro, Expressing, Control

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: GSTP1 inhibited cell proliferation and drug resistance in a wtp53‐dependent manner in vitro. (A) MTT assays in si1‐GSTP1 and sictrl transfected wtp53 Capan‐2 cells cultured within 4 days. (B) MTT assays of GFP, GSTP1‐GFP and GSTP1‐GFP plus si‐p53 transfected SW1990 cells with wtp53 cultured within 4 days. (C, D) MTT assays in si1‐GSTP1 and sictrl transfected wtp53 Capan‐2 cells under OXA (C) or GEM (D) treatment for 2 days. (E, F) MTT assays in GFP, GSTP1‐GFP and GSTP1‐GFP plus si‐p53 transfected SW1990 cells with wtp53 under OXA (E) or GEM (F) treatment for 2 days. The experiments were performed in triplicate. Error bars ± SDs. * p < 0.05 and ** p < 0.01 compared with the control.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: In Vitro, Transfection, Cell Culture, Control

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: GSTP1 regulated cell proliferation and drug resistance via p53/p21‐ and Bax/Bcl2‐mediated signaling pathways. (A, B) p53/p21 and Bax/Bcl2 protein expression in si1‐GSTP1 and sictrl transfected wtp53 Capan‐2 cells with or without OXA (A) or GEM (B) treatment for 2 days. (C, D) p53/p21 and Bax/Bcl2 protein expression in GFP and GSTP1‐GFP transfected SW1990 cells with wtp53 with or without OXA (C) or GEM (D) treatment for 2 days. 1: Sictrl group; 2: si1‐GSTP1 group; 3: Sictrl with OXA (A) or GEM (B) treatment group; 4: si1‐GSTP1 with OXA (A) or GEM (B) treatment group; 5: GFP group; 6: GSTP1‐GFP group; 7: GFP with OXA (C) or GEM (D) treatment group; 8: GSTP1‐GFP with OXA (C) or GEM (D) treatment group. The experiments were performed in triplicate. Error bars ± SDs. * p < 0.05 and ** p < 0.01 compared with the control.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: Protein-Protein interactions, Expressing, Transfection, Control

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: GSTP1 mediated cell invasion and migration via partially regulating wtp53 mediated EMT signaling. (A) Cell invasion and migration in sictrl, si1‐GSTP1 and si2‐GSTP1 transfected wtp53 Capan‐2 cells. (B) Cell invasion and migration of GFP, GSTP1‐GFP and GSTP1‐GFP plus si‐p53 transfected SW1990 cells with wtp53. (C) Protein expression of EMT epithelial and mesenchymal biomarkers in sictrl, si1‐GSTP1 and si2‐GSTP1 transfected wtp53 Capan‐2 cells. (D) Protein expression of EMT epithelial and mesenchymal biomarkers in GFP and GSTP1‐GFP transfected SW1990 cells with wtp53. 1: Sictrl group; 2: si1‐GSTP1 group; 3: si2‐GSTP1 group; 4: GFP group; 5: GSTP1‐GFP group. The experiments were performed in triplicate. Error bars ± SDs. * p < 0.05 and ** p < 0.01 compared with the control.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: Migration, Transfection, Expressing, Control

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: The feedback regulation of GSTP1 and wtp53 mediated the malignant progression of PC in vitro. (A) GSTP1 was not coimmunoprecipitated with p53 in mtp53 PANC‐1 or wtp53 Capan‐2 cells. (B) P53 protein could bind to the GSTP1 DNA promoter in wtp53 Capan‐2 or SW1990 cells. (C) P53 protein could not bind to the GSTP1 DNA promoter in mtp53 BxPC‐3 or PANC‐1 cells. (D) IF staining of p53 (TRITC, red) and nuclei (Hoechst, blue) in si1‐GSTP1 and sictrl transfected wtp53 Capan‐2 cells. (E) IF staining of p53 (TRITC, red) and nuclei (Hoechst, blue) in GFP and GSTP1‐GFP transfected SW1990 cells with wtp53. (F) IF staining of p53 (TRITC, red) and nuclei (Hoechst, blue) in si1‐GSTP1 and sictrl transfected mtp53 BxPC‐3 cells. (G) IF staining of p53 (TRITC, red) and nuclei (Hoechst, blue) in si1‐GSTP1 and sictrl transfected mtp53 PANC‐1 cells.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: In Vitro, Staining, Transfection

Journal: Cancer Science

Article Title: A Mutual Interaction Between GSTP1 and p53 Improves the Drug Resistance and Malignant Biology of Pancreatic Cancer

doi: 10.1111/cas.70019

Figure Lengend Snippet: GSTP1 inhibited tumor growth in coordination with wtp53 in vivo. (A) Subcutaneous tumor volumes in GFP and GSTP1‐GFP transfected SW1990 cells with wtp53 in 5 nude mice. (B) HE staining of the subcutaneous tumor. (C) GSTP1 and p53 expression in subcutaneous tumors in the GFP and GSTP1‐GFP groups, as determined by WB. (D) ki67 expression in the subcutaneous tumors of the GFP and GSTP1‐GFP groups was compared via IHC. (E) Liver metastases in GFP and GSTP1‐GFP transfected SW1990 cells with wtp53 in 5 nude mice. (F) HE staining of liver metastases. (G) Representative images from the IHC assays of GSTP1 and p53 protein expression in the GFP and GSTP1‐GFP groups in vivo. (H) Statistical data from the IHC assays of GSTP1 and p53 protein expression in the GFP and GSTP1‐GFP groups in vivo. Error bars ± SDs. * p < 0.05 and ** p < 0.01 compared with the control.

Article Snippet: The wtp53 Capan‐2 (Kras mutant) cell line was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) [ ].

Techniques: In Vivo, Transfection, Staining, Expressing, Control

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: LAI-1 and lysosomes subcellular colocalization in A549 cell line. Live cell imaging fluorescence microscopy was performed. ( A ) Differential interference contrast of A549 cells. ( B ) Lysotracker Green labelling lysosomes (green channel). ( C ) A total of 10 µM of LAI-1 treatment after 3 h (red channel). ( D ) Merged channel showing lysosomes and LAI-1 colocalization (orange). Images are representative of three independent experiments. Scale bar 30 µm. Colocalization between Lysotracker and LAI-1 was quantified using Pearson’s correlation coefficient (PCC) and Mander’s overlap coefficient (MOC).

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Live Cell Imaging, Fluorescence, Microscopy

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Cell viability dose–response curves, evaluated with MTT assay, for all cell lines (A549, SW900 and DMS53) after 24 h incubation with chloroquine (CQ), 3-Methyladenine (3-MA) and LAI-1 treatment at different concentrations. Each point represents the mean value ± SD.

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: MTT Assay, Incubation

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Inhibitory concentration (IC); IC 25 , IC 50 and IC 75 values after LAI-1 treatment for A549, SW900 and DMS53 cell lines. Data show mean value ± SD.

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Concentration Assay

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Autophagy markers after treatment with different autophagy inhibitors. ( A ) Western blot showing autophagy-related proteins LC3 and p62/SQSTM1 after 24 h with chloroquine (CQ, 150 µM), 3-Methyladenine (3-MA, 10 mM) and LAI-1 (10 µM) treatment in A549 cells. ( B ) LC3-I and LC3-II protein levels and their LC3-II/I ratios. ( C ) p62/SQSTM1 protein expression. Protein expression was normalized using GAPDH as loading control. Fold induction against control group (CT) was calculated. Figure shows mean ± SEM. Statistical differences against CT are shown as *** p < 0.001, ** p < 0.01 and * p < 0.05. The whole western blot figures are showed in .

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Western Blot, Expressing, Control

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Autophagy modulation after LAI-1 treatment. ( A ) Dose–response Western blot showing autophagy-related proteins LC3 and p62/SQSTM1 after 24 h of LAI-1 treatment at different concentrations (10, 15 and 20 µM) in A549, SW900 and DMS53 cell lines. ( B ) LC3-I and LC3-II protein levels and their LC3-II/I ratios after dose–response assay. ( C ) p62/SQSTM1 protein expression after dose–response experiment. ( D ) Time-course Western blot showing autophagy-related proteins LC3 and p62/SQSTM1 after treating A549 cells with LAI-1 (10 µM) at different time points (0, 4, 8, 16, 24, 48 h). ( E ) LC3-I and LC3-II protein levels and their LC3-II/I ratios after time-course experiment. ( F ) p62/SQSTM1 protein expression after time-course assay. ( G ) Time-course Western blot showing autophagy activation proteins Akt and mTOR after treating A549 cells with LAI-1 (10 µM) at different time points (0, 4, 8 h). ( H ) Akt and phospho-Akt at S473 (pAkt) protein levels after time-course experiment. ( I ) mTOR and phospho-mTOR at S2778 (pmTOR) protein expression after time-course assay. Protein expression was normalized using GAPDH as loading control. Fold induction against control group (CT) was calculated. Figure shows mean ± SEM. Statistical differences against CT are shown as * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001. The whole western blot figures are showed in .

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Western Blot, Expressing, Activation Assay, Control

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Subcellular localization of LC3 and LAMP-1. A549 cells were harvested for 24 h on coverslips and then treated with LAI-1 (15 µM) and chloroquine (CQ, 50 µM) for different times. The fusion between autophagosomes, marked with LC3 (green), and lysosomes, marked with LAMP-1 (red), was studied in merged images. Zoomed images were used to better observe this process. The localization and intensity of LC3 staining were also analyzed. DAPI (blue) staining was used for nuclear localization. Images are representative of three independent experiments. Scale bar 30 µm.

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Staining

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Lysosomal and intracellular pH modifications using LAI-1 treatment. ( A ) A549 cells were treated for 1 h with LAI-1 or chloroquine (CQ) at different concentrations. After that, acridine orange staining (5 µg/mL) was performed for 30 min at room temperature. Images are representative of three independent experiments. Scale bar 50 µm. ( B ) Intracellular pH measurement in A549 cells treated with different LAI-1 concentrations for 1 h. Staining with pH Rodo Red AM staining kit and a calibration curve was conducted to quantify intracellular pH. Figure shows mean ± SEM. Statistical differences against control group (CT) are shown as ** p < 0.01.

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Staining, Control

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Cell death characterization using flow cytometry. The different cell lines (A549, SW900 and DMS53) were treated for 24 h with different LAI-1 concentrations. Cells were stained with Annexin-V APC/ Sytox Green kit. Early apoptotic cells were defined as Annexin-positive and Sytox-negative, whereas late apoptotic or necrotic cells were defined as Annexin- and Sytox-positive. ( A ) Representative plots. ( B ) Quantification of % of cells in early apoptosis and in late apoptosis or necrosis after different treatment. Statistical differences against control group (CT) are shown as * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001.

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Flow Cytometry, Staining, Control

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Expression of apoptotic and cell-cycle related proteins. The different cell lines (A549, SW900 and DMS53) were treated for 24 h with different LAI-1 concentrations and several proteins were assessed with Western blot ( A ). Quantification of procaspase 3 ( B ), PARP ( C ), cleaved-PARP ( D ), p21 ( E ) and p53 ( F ) protein expression after Western blot analysis was performed using GAPDH expression as loading control. Fold induction against control group (CT) was calculated. Figures show mean ± SEM. Statistical differences against CT are shown as * p < 0.05, ** p < 0.01 and *** p < 0.001. The whole western blot figures are showed in .

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: Expressing, Western Blot, Control

Journal: Cancers

Article Title: A Novel Late-Stage Autophagy Inhibitor That Efficiently Targets Lysosomes Inducing Potent Cytotoxic and Sensitizing Effects in Lung Cancer

doi: 10.3390/cancers14143387

Figure Lengend Snippet: Cell viability, evaluated with MTT assay, for A549, SW900 and DMS53 lung cancer cell lines after 24 h incubation with cisplatin (CisPt, 40 µM), LAI-1 (15 µM) and the combination. Figure shows mean ± SEM. Statistical differences are shown as **** p < 0.0001 ** p < 0.01 and * p < 0.05.

Article Snippet: Human lung cancer cell lines A549 (adenocarcinoma, KRAS-mutated), DMS53 (small cell carcinoma, p53-mutated) and SW900 (squamous carcinoma, KRAS- and p53-mutated) were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in DMEM (A549) or RPMI (DMS53 and SW900) medium (Biological Industries, Beit Haemek, Israel).

Techniques: MTT Assay, Incubation