docetaxel Search Results


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  • 99
    Millipore docetaxel
    Induction of cellular apoptosis by EL102 and <t>docetaxel.</t> Apoptosis in ( A ) CWR22, ( B ) 22Rv1, ( C ) PC-3 and ( D ) DU145 prostate cancer cell lines as measured by the percentage of cells accumulated in subG 1 . (See Supplementary Table 4 for results of one-way ANOVA comparing cells in subG 1 /apoptosis between each treatment). ( E ) PARP cleavage in DU145 protein lysates 24 and 48 h post treatment with EL102 and docetaxel by western blot. Abbreviation: DCT=docetaxel.
    Docetaxel, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 511 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    Sanofi docetaxel
    Effects of <t>docetaxel</t> on ( A ) SKOV3 and SKOV3-TR or ( B ) HeyA8 and HeyA8-MDR ovarian cancer cells. The percentage of apoptosis was determined by terminal deoxynucleotidyl transferase – mediated nick end labeling. Cells were treated with or without
    Docetaxel, supplied by Sanofi, used in various techniques. Bioz Stars score: 95/100, based on 382 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    LC Laboratories docetaxel
    Effect of exogenous expression of MRP2 on the cytotoxicity ( a ) and intracellular accumulation ( b ) of <t>docetaxel.</t> ( a ) Vector-transfected MDCKII cells (closed circles) or MRP2-expressing MDCKII cells (open circles) were incubated for 96 h with various concentrations (0.01–100 nmol/l) of docetaxel. Cell survival was quantified by the MTT assay as described in the Methods. ( b ) Cell lines were incubated for 24 h with 1 nmol/l of docetaxel, and the volume uptake of docetaxel into these cells was determined. Each point and bar represents the mean ± SE ( n = 3). * P
    Docetaxel, supplied by LC Laboratories, used in various techniques. Bioz Stars score: 94/100, based on 118 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Selleck Chemicals docetaxel
    Forkhead box protein M1 (FOXM1) promotes <t>docetaxel</t> resistance through regulating the microtubule-destabilizing protein Stathmin rather than mitotic centromere–associated kinesin (MCAK) in gastric cancers. ( A ) mRNA levels of FOXM1, Stathmin and MCAK in AGS cells after transfected with pcDNA3. 1, pcDNA3. 1-FOXM1, non-specific siRNA or FOXM1-siRNA were determined by RT-PCR. ( B ) The protein expression levels of FOXM1, Stathmin and MCAK after transfections were shown by western blot analysis. ( C ) Polymerized and soluble tubulin fractions from siRNA -FOXM1, siRNA-Stathmin, siRNA-MCAK and non-specific siRNA-transfected AGS-DOC R cell lines treated or untreated with docetaxel were detected by western blot (Left). The relative polymerized microtubule fractions of both α-tubulin and β-tubulin were significantly increased after the treatment of docetaxel in FOXM1, Stathmin and MCAK knockdown AGS-DOC R cells (Right). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.
    Docetaxel, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 93/100, based on 66 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Enzo Biochem docetaxel
    <t>Docetaxel</t> induces enlargement and expansion of tumor-draining lymphatics in vivo that can be attenuated by VEGFR3 blockade. a Quantified lymphatic vessel perimeter in both tumor-bearing mammary fat pad of treated 4T1 mice and contralateral naïve (non-tumor bearing) fat pad. b Quantified lymphatic vessel area in both tumor-bearing mammary fat pad of treated 4T1 mice and contralateral naïve (non-tumor bearing) fat pad. c Quantified lymphatic vessel density of single podoplanin+ lymphatic vessels per stromal area throughout sections. d Total lymphatic endothelial cells per weight of fat pad (live CD45 − CD31 + gp38 + ) as determined by flow cytometry. e Fluid drainage from tumor to axillary lymph node as determined after Evans blue injection into tumor-bearing mammary fat pad 24 h after treatment with docetaxel. ( n = 5–9 mice per group). f Quantified lymphatic vessel density displayed as number of peritumoral lymphatic vessels per mm 2 of stromal tissue after 0, 1, 2, and 3 doses of docetaxel (8 mg/kg, IV, 3 days apart). * p
    Docetaxel, supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 93/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Aventis docetaxel
    Effects of <t>docetaxel</t> on ( A ) SKOV3 and SKOV3-TR or ( B ) HeyA8 and HeyA8-MDR ovarian cancer cells. The percentage of apoptosis was determined by terminal deoxynucleotidyl transferase – mediated nick end labeling. Cells were treated with or without
    Docetaxel, supplied by Aventis, used in various techniques. Bioz Stars score: 92/100, based on 138 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Jiangsu Hengrui Medicine docetaxel
    Bax protein expression induced by 0 nM and 20 nM <t>docetaxel</t> in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 (A) and H460 (B) cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, Western blot analyses were performed to detect the expression of Bax. The graph depicts mean±SEM for three independent determinations of optical density of the Bax Western blot bands. * Compared to control cells with incubation of 0 nM docetaxel, p
    Docetaxel, supplied by Jiangsu Hengrui Medicine, used in various techniques. Bioz Stars score: 89/100, based on 49 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    77
    Green Plantchem Company Limited drug docetaxel
    Decay-corrected TACs of [ 11 <t>C]docetaxel</t> (%ID/ml versus time p.i.) in plasma and whole blood
    Drug Docetaxel, supplied by Green Plantchem Company Limited, used in various techniques. Bioz Stars score: 77/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    76
    FUJIFILM docetaxel trihydrate
    Typical mass spectra obtained from a the spot of the <t>docetaxel</t> using ( a ) α-cyano-4-hydroxycinnamic acid (CHCA); ( b ) 2,5-dihydroxy benzoic acid (DHB); ( c ) 4-nitroaniline (4NA); and ( d ) 6-aza-2-thiothymine (ATT). M indicates docetaxel.
    Docetaxel Trihydrate, supplied by FUJIFILM, used in various techniques. Bioz Stars score: 76/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Induction of cellular apoptosis by EL102 and docetaxel. Apoptosis in ( A ) CWR22, ( B ) 22Rv1, ( C ) PC-3 and ( D ) DU145 prostate cancer cell lines as measured by the percentage of cells accumulated in subG 1 . (See Supplementary Table 4 for results of one-way ANOVA comparing cells in subG 1 /apoptosis between each treatment). ( E ) PARP cleavage in DU145 protein lysates 24 and 48 h post treatment with EL102 and docetaxel by western blot. Abbreviation: DCT=docetaxel.

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Induction of cellular apoptosis by EL102 and docetaxel. Apoptosis in ( A ) CWR22, ( B ) 22Rv1, ( C ) PC-3 and ( D ) DU145 prostate cancer cell lines as measured by the percentage of cells accumulated in subG 1 . (See Supplementary Table 4 for results of one-way ANOVA comparing cells in subG 1 /apoptosis between each treatment). ( E ) PARP cleavage in DU145 protein lysates 24 and 48 h post treatment with EL102 and docetaxel by western blot. Abbreviation: DCT=docetaxel.

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques: Western Blot

    Cell cycle analysis of DU145 cell accumulation in G1, S, G2/M and subG 1 after EL102, docetaxel or combination treatment. Cell cycle profiles of DU145 cells, with markers indicating the G1, S, G2/M and subG 1 cells, after exposure to ( A ) 10 n M EL102 or 1 n M docetaxel, or 10 n M EL102 and 1 n M docetaxel, ( B ) 10 n M EL102 or 10 n M docetaxel, or 10 n M EL102 and 10 n M docetaxel, ( C ) 100 n M EL102 or 1 n M docetaxel, or 100 n M EL102 and 1 n M docetaxel, ( D ) 100 n M EL102 or 10 n M docetaxel, or 100 n M EL102 and 10 n M docetaxel for 24, 48 and 72 h. Graphs of cell cycle analysis of accumulation of DU145 in the G 1 , S, G 2 /M and sub-G 1 phase in response to 0, 10 and 100 n M EL102, 1 and 10 n M docetaxel and combinations of each at ( E ) 24, ( F ) 48, ( G ) 72 h as measured by propidium iodide flow cytometry.

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Cell cycle analysis of DU145 cell accumulation in G1, S, G2/M and subG 1 after EL102, docetaxel or combination treatment. Cell cycle profiles of DU145 cells, with markers indicating the G1, S, G2/M and subG 1 cells, after exposure to ( A ) 10 n M EL102 or 1 n M docetaxel, or 10 n M EL102 and 1 n M docetaxel, ( B ) 10 n M EL102 or 10 n M docetaxel, or 10 n M EL102 and 10 n M docetaxel, ( C ) 100 n M EL102 or 1 n M docetaxel, or 100 n M EL102 and 1 n M docetaxel, ( D ) 100 n M EL102 or 10 n M docetaxel, or 100 n M EL102 and 10 n M docetaxel for 24, 48 and 72 h. Graphs of cell cycle analysis of accumulation of DU145 in the G 1 , S, G 2 /M and sub-G 1 phase in response to 0, 10 and 100 n M EL102, 1 and 10 n M docetaxel and combinations of each at ( E ) 24, ( F ) 48, ( G ) 72 h as measured by propidium iodide flow cytometry.

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques: Cell Cycle Assay, Flow Cytometry, Cytometry

    Impact of EL102 and docetaxel alone and in combination on CWR22 xenograft tumour volume. ( A ) Effect of vehicle vs 12 mg kg −1 docetaxel, vs 12 mg kg −1 EL102, vs 15 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 12 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 15 mg kg −1 EL102, on CWR22 tumour volume using a 5-day on/2-day off schedule (tumour volume (cm 3 )± s.e.m.). (See Supplementary Table 1 for one-way ANOVA comparing tumour volume at each time point). ( B ) Effect of vehicle vs 12 mg kg −1 docetaxel, vs 12 mg kg −1 EL102, vs 15 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 12 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 15 mg kg −1 EL102, on mouse body weight (Note: vehicle group killed on day 24 due to tumour size). (See Supplementary Table 2 for one-way ANOVA comparing body weight at each time point.)

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Impact of EL102 and docetaxel alone and in combination on CWR22 xenograft tumour volume. ( A ) Effect of vehicle vs 12 mg kg −1 docetaxel, vs 12 mg kg −1 EL102, vs 15 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 12 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 15 mg kg −1 EL102, on CWR22 tumour volume using a 5-day on/2-day off schedule (tumour volume (cm 3 )± s.e.m.). (See Supplementary Table 1 for one-way ANOVA comparing tumour volume at each time point). ( B ) Effect of vehicle vs 12 mg kg −1 docetaxel, vs 12 mg kg −1 EL102, vs 15 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 12 mg kg −1 EL102, vs 12 mg kg −1 docetaxel plus 15 mg kg −1 EL102, on mouse body weight (Note: vehicle group killed on day 24 due to tumour size). (See Supplementary Table 2 for one-way ANOVA comparing body weight at each time point.)

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques:

    Effect of EL102 on microtubule destabilisation. The effects of EL102 treatments (0, 10, 100 n M ), docetaxel treatments (0, 1 and 10 n M ) and a combination of both, on microtubule stability in DU145 prostate cancer cells were determined. Following 24 h post-treatment incubation at optimum conditions, cells were fixed with ice-cold methanol and simultaneously stained with Rabbit anti- β -tubulin and anti-acetylated tubulin antibodies. Microtubules were visualised using Alexa Fluor 647 donkey anti-rabbit IgG secondary and Rhodamine Red-X-AffiniPure Fab Fragment goat anti-mouse IgG. Nuclei were counterstained using DAPI.

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Effect of EL102 on microtubule destabilisation. The effects of EL102 treatments (0, 10, 100 n M ), docetaxel treatments (0, 1 and 10 n M ) and a combination of both, on microtubule stability in DU145 prostate cancer cells were determined. Following 24 h post-treatment incubation at optimum conditions, cells were fixed with ice-cold methanol and simultaneously stained with Rabbit anti- β -tubulin and anti-acetylated tubulin antibodies. Microtubules were visualised using Alexa Fluor 647 donkey anti-rabbit IgG secondary and Rhodamine Red-X-AffiniPure Fab Fragment goat anti-mouse IgG. Nuclei were counterstained using DAPI.

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques: Incubation, Staining

    Impact of EL102 and docetaxel combination treatment on prostate cancer cell line viability in vitro . Effect of EL102 and docetaxel in combination of in vitro cell viability after 72 h in ( A ) CWR22, ( B ) 22Rv1, ( C ) PC-3 and ( D ) DU145 prostate cancer cell. (See Supplementary Table 3 for results of one-way ANOVA comparing cell viability between each treatment.)

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Impact of EL102 and docetaxel combination treatment on prostate cancer cell line viability in vitro . Effect of EL102 and docetaxel in combination of in vitro cell viability after 72 h in ( A ) CWR22, ( B ) 22Rv1, ( C ) PC-3 and ( D ) DU145 prostate cancer cell. (See Supplementary Table 3 for results of one-way ANOVA comparing cell viability between each treatment.)

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques: In Vitro

    Impact of EL102 and docetaxel on prostate cancer cell line viability in vitro . ( A ) Chemical structure of EL102. ( B ) Dose response effects of EL102 on prostate cancer cell line viability over 72-h exposure. ( C ) Dose response effects of docetaxel on prostate cancer cell line viability over 72-h exposure. ( D ) Effect of EL102 on doxorubicin and docetaxel-resistant DLKPA lung cancer cell line viability vs DLKP parental lung cancer cell line. ( E ) Comparison of docetaxel sensitivity in the doxorubicin and docetaxel-resistant DLKPA lung cancer cell line viability vs DLKP parental lung cancer cell line.

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Impact of EL102 and docetaxel on prostate cancer cell line viability in vitro . ( A ) Chemical structure of EL102. ( B ) Dose response effects of EL102 on prostate cancer cell line viability over 72-h exposure. ( C ) Dose response effects of docetaxel on prostate cancer cell line viability over 72-h exposure. ( D ) Effect of EL102 on doxorubicin and docetaxel-resistant DLKPA lung cancer cell line viability vs DLKP parental lung cancer cell line. ( E ) Comparison of docetaxel sensitivity in the doxorubicin and docetaxel-resistant DLKPA lung cancer cell line viability vs DLKP parental lung cancer cell line.

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques: In Vitro

    Impact of EL102 and docetaxel alone and in combination on tubulin polymerisation activity. The change in OD ± s.e.m over time (mins) was measured following tubulin's treatment with 5 μ M EL102, 2 μ M docetaxel and a combination of both vs untreated and 2 μ M nocodazole.

    Journal: British Journal of Cancer

    Article Title: The novel toluidine sulphonamide EL102 shows pre-clinical in vitro and in vivo activity against prostate cancer and circumvents MDR1 resistance

    doi: 10.1038/bjc.2013.537

    Figure Lengend Snippet: Impact of EL102 and docetaxel alone and in combination on tubulin polymerisation activity. The change in OD ± s.e.m over time (mins) was measured following tubulin's treatment with 5 μ M EL102, 2 μ M docetaxel and a combination of both vs untreated and 2 μ M nocodazole.

    Article Snippet: Docetaxel was purchased from Sigma-Aldrich (Dublin, Ireland; #01885-5MG-F).

    Techniques: Activity Assay

    Determination of scAb on the nanoparticle surface. Confocal microscopic images of coumarin(green)-loaded scAb-PLGA-SPIO/coumarin ( A ) and PLGA-SPIO/coumarin ( B ) incubated with PE(red)-labeled secondary antibodies. The merged fluorescence indicated by the arrows demonstrates that scAb was successfully conjugated to the nanoparticle surface. ( C ) A significant shift of PE fluorescence intensity could be observed for scAb-PLGA-SPIO/docetaxel (scAb-NPs) in comparison with blank control and PLGA-SPIO/docetaxel NPs, indicating the presence of scAb on the nanoparticle surface. Abbreviations: NPs, nanoparticles; PLGA, poly(D,L-lactic-co-glycolic acid); SPIO, superparamagnetic iron oxide; PE, phycoerythrin.

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: Determination of scAb on the nanoparticle surface. Confocal microscopic images of coumarin(green)-loaded scAb-PLGA-SPIO/coumarin ( A ) and PLGA-SPIO/coumarin ( B ) incubated with PE(red)-labeled secondary antibodies. The merged fluorescence indicated by the arrows demonstrates that scAb was successfully conjugated to the nanoparticle surface. ( C ) A significant shift of PE fluorescence intensity could be observed for scAb-PLGA-SPIO/docetaxel (scAb-NPs) in comparison with blank control and PLGA-SPIO/docetaxel NPs, indicating the presence of scAb on the nanoparticle surface. Abbreviations: NPs, nanoparticles; PLGA, poly(D,L-lactic-co-glycolic acid); SPIO, superparamagnetic iron oxide; PE, phycoerythrin.

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Incubation, Labeling, Fluorescence

    ( A ) Confocal laser scanning microscopy of PC3M cells (prostate stem cell antigen-positive) and HT29 cells (prostate stem cell antigen-negative) after 2 hours of incubation with void nanoparticles (control), PLGA-SPIO/coumarin, and scAb-PLGA-SPIO/coumarin, respectively. ( B ) Mean fluorescence intensity of PC3M and HT29 cells in Figure 5A . ( C ) In vitro T2-weighted images (cross section and sagittal section) of untreated PC3M cells, PC3M cells incubated with scAb-PLGA-SPIO/docetaxel or PLGA-SPIO/docetaxel, and HT29 cells incubated with scAb-PLGA-SPIO/docetaxel. ( D ) T 2 values of PC3M and HT29 cells described in Figure 5C . Notes: The results are shown as the mean ± standard error of the mean (n = 3). * P

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: ( A ) Confocal laser scanning microscopy of PC3M cells (prostate stem cell antigen-positive) and HT29 cells (prostate stem cell antigen-negative) after 2 hours of incubation with void nanoparticles (control), PLGA-SPIO/coumarin, and scAb-PLGA-SPIO/coumarin, respectively. ( B ) Mean fluorescence intensity of PC3M and HT29 cells in Figure 5A . ( C ) In vitro T2-weighted images (cross section and sagittal section) of untreated PC3M cells, PC3M cells incubated with scAb-PLGA-SPIO/docetaxel or PLGA-SPIO/docetaxel, and HT29 cells incubated with scAb-PLGA-SPIO/docetaxel. ( D ) T 2 values of PC3M and HT29 cells described in Figure 5C . Notes: The results are shown as the mean ± standard error of the mean (n = 3). * P

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Confocal Laser Scanning Microscopy, Incubation, Fluorescence, In Vitro

    ( A ) IC 50 of PC3M (prostate stem cell antigen-positive) cells after 24, 48, and 72 hours of incubation with docetaxel, PLGA/docetaxel, scAb-PLGA/docetaxel, PLGA-SPIO/docetaxel, or scAb-PLGA-SPIO/docetaxel. The Cell Counting Kit-8 assay was used to evaluate cytotoxicity. The data are expressed as the mean ± standard error of the mean (n = 3). *,# P

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: ( A ) IC 50 of PC3M (prostate stem cell antigen-positive) cells after 24, 48, and 72 hours of incubation with docetaxel, PLGA/docetaxel, scAb-PLGA/docetaxel, PLGA-SPIO/docetaxel, or scAb-PLGA-SPIO/docetaxel. The Cell Counting Kit-8 assay was used to evaluate cytotoxicity. The data are expressed as the mean ± standard error of the mean (n = 3). *,# P

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Incubation, Cell Counting

    scAb-PLGA-SPIO/docetaxel demonstrated a superior outcome in a PC3M xenograft animal model. ( A ) Volumes of PC3M carcinoma in different groups. ( B ) Representative PC3M tumors excised from different groups. ( C ) Kaplan–Meier survival curve demonstrates that scAb-PLGA-SPIO/docetaxel significantly increased the lifespan of the mice. ( D ) Hematoxylin-eosin staining and terminal transferase uridyl nick-end labeling (TUNEL) staining of excised median tumors at 100× magnification. Histological staining was evaluated by two independent pathologists. Nuclei with dark brown horseradish peroxidase staining indicates apoptosis. ( E ) Body weight of the nude mice in different groups. ( F ) Effect of different treatments on the white blood cell counts at the endpoint of observation. Note: The results are shown as the mean ± standard error of the mean (n = 5). Abbreviations: Dtxl, docetaxel; PLGA, poly(D,L-lactic-co-glycolic acid); SPIO, superparamagnetic iron oxide; PBS, phosphate-buffered solution.

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: scAb-PLGA-SPIO/docetaxel demonstrated a superior outcome in a PC3M xenograft animal model. ( A ) Volumes of PC3M carcinoma in different groups. ( B ) Representative PC3M tumors excised from different groups. ( C ) Kaplan–Meier survival curve demonstrates that scAb-PLGA-SPIO/docetaxel significantly increased the lifespan of the mice. ( D ) Hematoxylin-eosin staining and terminal transferase uridyl nick-end labeling (TUNEL) staining of excised median tumors at 100× magnification. Histological staining was evaluated by two independent pathologists. Nuclei with dark brown horseradish peroxidase staining indicates apoptosis. ( E ) Body weight of the nude mice in different groups. ( F ) Effect of different treatments on the white blood cell counts at the endpoint of observation. Note: The results are shown as the mean ± standard error of the mean (n = 5). Abbreviations: Dtxl, docetaxel; PLGA, poly(D,L-lactic-co-glycolic acid); SPIO, superparamagnetic iron oxide; PBS, phosphate-buffered solution.

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Animal Model, Mouse Assay, Staining, End Labeling, TUNEL Assay

    In vivo assessment of dual-function nanoparticles as a negative MRI contrast enhancement agent. ( A ) Representative MRI coronal contrast images of mice bearing PC3M xenografts prior to injection and at various intervals after systemic administration of scAb-PLGA-SPIO/docetaxel or PLGA-SPIO/docetaxel via the tail vein. The arrows indicate the location of the tumors. ( B ) Normalized MRI signal intensities of the PC3M xenografts in Figure 6A . Notes: The results are shown as the mean ± standard error of the mean (n = 3). * P

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: In vivo assessment of dual-function nanoparticles as a negative MRI contrast enhancement agent. ( A ) Representative MRI coronal contrast images of mice bearing PC3M xenografts prior to injection and at various intervals after systemic administration of scAb-PLGA-SPIO/docetaxel or PLGA-SPIO/docetaxel via the tail vein. The arrows indicate the location of the tumors. ( B ) Normalized MRI signal intensities of the PC3M xenografts in Figure 6A . Notes: The results are shown as the mean ± standard error of the mean (n = 3). * P

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: In Vivo, Magnetic Resonance Imaging, Mouse Assay, Injection

    ( A ) Dynamic light scattering histogram showing the size distribution of scAb-PLGA-SPIO/docetaxel. ( B ) Transmission electron microscopic images of scAb-PLGA-SPIO/docetaxel. The nanoparticles were indicated by black arrows, and SPIO nanoparticles are shown with white arrows. ( C ) T 2 -weighted magnetic resonance images (repetition time 5000 msec, echo time 100 msec) of scAb-PLGA-SPIO/docetaxel at various iron concentrations. ( D ) T 2 relaxation rate as a function of the iron concentration of scAb-PLGA-SPIO/docetaxel. ( E ) In vitro docetaxel release profiles indicate that scAb-PLGA-SPIO/docetaxel showed slower drug release than scAb-PLGA/docetaxel. Note: Results are shown as the mean ± standard error of the mean (n = 3). Abbreviations: Dtxl, docetaxel; PLGA, poly(D,L-lactic-co-glycolic acid); SPIO, superparamagnetic iron oxide.

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: ( A ) Dynamic light scattering histogram showing the size distribution of scAb-PLGA-SPIO/docetaxel. ( B ) Transmission electron microscopic images of scAb-PLGA-SPIO/docetaxel. The nanoparticles were indicated by black arrows, and SPIO nanoparticles are shown with white arrows. ( C ) T 2 -weighted magnetic resonance images (repetition time 5000 msec, echo time 100 msec) of scAb-PLGA-SPIO/docetaxel at various iron concentrations. ( D ) T 2 relaxation rate as a function of the iron concentration of scAb-PLGA-SPIO/docetaxel. ( E ) In vitro docetaxel release profiles indicate that scAb-PLGA-SPIO/docetaxel showed slower drug release than scAb-PLGA/docetaxel. Note: Results are shown as the mean ± standard error of the mean (n = 3). Abbreviations: Dtxl, docetaxel; PLGA, poly(D,L-lactic-co-glycolic acid); SPIO, superparamagnetic iron oxide.

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Transmission Assay, Concentration Assay, In Vitro

    Synthetic scheme and preparation process for scAb-PLGA-SPIO/docetaxel using layer-by-layer strategy. Abbreviations: PEG, poly(ethylene glycol); EDC, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide; NHS, N-hydroxysuccinimide; PAH, poly(allylamine hydrochloride); PLGA, poly(D,L-lactic-co-glycolic acid); SPION, superparamagnetic iron oxide nanoparticles; Dtxl, docetaxel.

    Journal: International Journal of Nanomedicine

    Article Title: Prostate stem cell antigen-targeted nanoparticles with dual functional properties: in vivo imaging and cancer chemotherapy

    doi: 10.2147/IJN.S32804

    Figure Lengend Snippet: Synthetic scheme and preparation process for scAb-PLGA-SPIO/docetaxel using layer-by-layer strategy. Abbreviations: PEG, poly(ethylene glycol); EDC, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide; NHS, N-hydroxysuccinimide; PAH, poly(allylamine hydrochloride); PLGA, poly(D,L-lactic-co-glycolic acid); SPION, superparamagnetic iron oxide nanoparticles; Dtxl, docetaxel.

    Article Snippet: Docetaxel, poly(allylamine hydrochloride) (PAH), and the other chemicals were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques:

    Western blot analysis of PARP cleavage in B88 cells treated with the combination of docetaxel (0.5 nM) and γ-tocotrienol (50 μM) for 48 h. Whole-cell fractions extracted from the cells were subjected to analysis. The molecular weights of uncleaved and cleaved PARP were 116 and 89 kDa, respectively. Results are representative of 3 independent experiments.

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: Western blot analysis of PARP cleavage in B88 cells treated with the combination of docetaxel (0.5 nM) and γ-tocotrienol (50 μM) for 48 h. Whole-cell fractions extracted from the cells were subjected to analysis. The molecular weights of uncleaved and cleaved PARP were 116 and 89 kDa, respectively. Results are representative of 3 independent experiments.

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: Western Blot

    (A) Effects of docetaxel and γ-tocotrienol on the NF-κB DNA binding activity in B88 cells. Cells were treated with docetaxel (0.5 nM), γ-tocotrienol (50 μM) and docetaxel (0.5 nM) + γ-tocotrienol (50 μM) for 24 h, and nuclear extracts were prepared and analyzed by EMSA with the κB site from the κ-light-chain enhanced in B cells. The docetaxel treatment of cells stimulated the constitutive NF-κB activity, while γ-tocotrienol inhibited constitutive NF-κB activity in B88 cells. Moreover, simultaneous treatment with both agents greatly suppressed the docetaxel-induced NF-κB activity in the cells. The specificity of the complex was analyzed by incubation with an excess (100-fold) of unlabeled κB oligonucleotides (competitor). (B) Effect of docetaxel (0.5 nM) on the expression of p65 and IκB-α proteins in B88 cells. Nuclear and cytoplasmic extracts were prepared from cells following docetaxel treatment for the indicated time points and analyzed by western blot analysis. The expression of nuclear p65 was augmented by docetaxel at 12, 24 and 48 h after treatment, and cytoplasmic IκB-α expression was conversely inhibited at these time-points. (C) Effect of γ-tocotrienol (50 μM) on the expression of p65 and IκB-α proteins in B88 cells. The expression of nuclear p65 protein decreased at 12 and 24 h after treatment, and cytoplasmic IκB-α expression was conversely enhanced at these time-points. (D) When the cells were treated with both agents, a significant decrease in nuclear p65 protein expression was detected in a time-dependent manner, whereas cytoplasmic IκB-α protein expression was similarly enhanced in a time-dependent manner.

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: (A) Effects of docetaxel and γ-tocotrienol on the NF-κB DNA binding activity in B88 cells. Cells were treated with docetaxel (0.5 nM), γ-tocotrienol (50 μM) and docetaxel (0.5 nM) + γ-tocotrienol (50 μM) for 24 h, and nuclear extracts were prepared and analyzed by EMSA with the κB site from the κ-light-chain enhanced in B cells. The docetaxel treatment of cells stimulated the constitutive NF-κB activity, while γ-tocotrienol inhibited constitutive NF-κB activity in B88 cells. Moreover, simultaneous treatment with both agents greatly suppressed the docetaxel-induced NF-κB activity in the cells. The specificity of the complex was analyzed by incubation with an excess (100-fold) of unlabeled κB oligonucleotides (competitor). (B) Effect of docetaxel (0.5 nM) on the expression of p65 and IκB-α proteins in B88 cells. Nuclear and cytoplasmic extracts were prepared from cells following docetaxel treatment for the indicated time points and analyzed by western blot analysis. The expression of nuclear p65 was augmented by docetaxel at 12, 24 and 48 h after treatment, and cytoplasmic IκB-α expression was conversely inhibited at these time-points. (C) Effect of γ-tocotrienol (50 μM) on the expression of p65 and IκB-α proteins in B88 cells. The expression of nuclear p65 protein decreased at 12 and 24 h after treatment, and cytoplasmic IκB-α expression was conversely enhanced at these time-points. (D) When the cells were treated with both agents, a significant decrease in nuclear p65 protein expression was detected in a time-dependent manner, whereas cytoplasmic IκB-α protein expression was similarly enhanced in a time-dependent manner.

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: Binding Assay, Activity Assay, Incubation, Expressing, Western Blot

    (A) In vitro morphogenetic behavior of cells (a) untreated or those treated with either (b) docetaxel (0.5 nM), (c) γ-tocotrienol (50 μM), or (d) both agents for 24 h. (a-d) A photomicrograph of Hoechst 33258-stained B88 nuclei revealed the chromatin condensation and nuclear fragmentation (red arrow heads) typical of apoptosis. (Original magnification, ×100.) (B) Effect of docetaxel (0.5 nM) and γ-tocotrienol (50 μM) on apoptotic cell death. Data represent the mean apoptotic cell death ± SD as follows: untreated control cells, 3.0±2.1%; docetaxel (0.5 nM)-treated cells, 7.9±3.1%; γ-tocotrienol (50 μM)-treated cells, 8.2±2.0%; and docetaxel (0.5 nM)- and γ-tocotrienol (50 μM)-treated cells, 55.1±5.2%. Results were analyzed by the Mann-Whitney U test. P

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: (A) In vitro morphogenetic behavior of cells (a) untreated or those treated with either (b) docetaxel (0.5 nM), (c) γ-tocotrienol (50 μM), or (d) both agents for 24 h. (a-d) A photomicrograph of Hoechst 33258-stained B88 nuclei revealed the chromatin condensation and nuclear fragmentation (red arrow heads) typical of apoptosis. (Original magnification, ×100.) (B) Effect of docetaxel (0.5 nM) and γ-tocotrienol (50 μM) on apoptotic cell death. Data represent the mean apoptotic cell death ± SD as follows: untreated control cells, 3.0±2.1%; docetaxel (0.5 nM)-treated cells, 7.9±3.1%; γ-tocotrienol (50 μM)-treated cells, 8.2±2.0%; and docetaxel (0.5 nM)- and γ-tocotrienol (50 μM)-treated cells, 55.1±5.2%. Results were analyzed by the Mann-Whitney U test. P

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: In Vitro, Staining, MANN-WHITNEY

    Growth suppression of human oral cancer (B88) cells by (A) docetaxel, (B) γ-tocotrienol and (C) docetaxel + γ-tocotrienol. The cells (5×10 3 cells per well) were grown in 96-well plates in medium supplemented with docetaxel [0.5 (▪), 1 (▴) and 2 nM (•)], γ-tocotrienol [50 (▪), 75 (▴) and 100 μM (•)], or docetaxel (0.5 nM) + γ-tocotrienol (50 μM) (•) for 6 days. Viable cells were estimated by MTT assay. The absorbance was measured at 570 nm. Standard deviations were calculated from 3 independent experiments. Growth was significantly lower compared to untreated or cells treated with lower concentrations of each agent. ★, Statistically significant at P

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: Growth suppression of human oral cancer (B88) cells by (A) docetaxel, (B) γ-tocotrienol and (C) docetaxel + γ-tocotrienol. The cells (5×10 3 cells per well) were grown in 96-well plates in medium supplemented with docetaxel [0.5 (▪), 1 (▴) and 2 nM (•)], γ-tocotrienol [50 (▪), 75 (▴) and 100 μM (•)], or docetaxel (0.5 nM) + γ-tocotrienol (50 μM) (•) for 6 days. Viable cells were estimated by MTT assay. The absorbance was measured at 570 nm. Standard deviations were calculated from 3 independent experiments. Growth was significantly lower compared to untreated or cells treated with lower concentrations of each agent. ★, Statistically significant at P

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: MTT Assay

    Induction of processing of initiator caspases and effector caspase by docetaxel and γ-tocotrienol. (A) Induction of caspase-3 processing by agents in B88 cells. The activation of caspase-3 was clearly observed at 24 and 48 h after treatment. Results are representative of 3 independent experiments. (B) Activation of caspase-8, as detected by cleavage of procaspase-8, was also evident after treatment with these agents. Results are representative of 3 independent experiments. (C) Induction of caspase-9 processing by agents. Docetaxel and γ-tocotrienol induced the activation of caspase-9. Results are representative of 3 independent experiments.

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: Induction of processing of initiator caspases and effector caspase by docetaxel and γ-tocotrienol. (A) Induction of caspase-3 processing by agents in B88 cells. The activation of caspase-3 was clearly observed at 24 and 48 h after treatment. Results are representative of 3 independent experiments. (B) Activation of caspase-8, as detected by cleavage of procaspase-8, was also evident after treatment with these agents. Results are representative of 3 independent experiments. (C) Induction of caspase-9 processing by agents. Docetaxel and γ-tocotrienol induced the activation of caspase-9. Results are representative of 3 independent experiments.

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: Activation Assay

    Western blot analysis for cytochrome c and Apaf-1 expression in cytosolic extracts in B88 cells. Cells were treated with docetaxel (0.5 nM) and γ-tocotrienol (50 μM) for the indicated periods of time, and cytosolic extracts were subjected to analysis. Both agents induced the release of cytochrome c into the cytoplasm. To ensure the lack of mitochondrial contamination of the extracts, the membrane was also probed with an antibody against mitochondrial cytochrome c oxidase subunit IV (data not shown). The treatment of cells with docetaxel and γ-tocotrienol also induced the expression of Apaf-1. As a loading control for the protein samples, the expression of β-actin is demonstrated. Results are representative of 3 independent experiments.

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: Western blot analysis for cytochrome c and Apaf-1 expression in cytosolic extracts in B88 cells. Cells were treated with docetaxel (0.5 nM) and γ-tocotrienol (50 μM) for the indicated periods of time, and cytosolic extracts were subjected to analysis. Both agents induced the release of cytochrome c into the cytoplasm. To ensure the lack of mitochondrial contamination of the extracts, the membrane was also probed with an antibody against mitochondrial cytochrome c oxidase subunit IV (data not shown). The treatment of cells with docetaxel and γ-tocotrienol also induced the expression of Apaf-1. As a loading control for the protein samples, the expression of β-actin is demonstrated. Results are representative of 3 independent experiments.

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: Western Blot, Expressing

    γ-tocotrienol inhibition of the anti-apoptotic gene products, survivin, cIAP-1, cIAP-2, XIAP and Bcl-2. Cells were left untreated or incubated with either (A) docetaxel (0.5 nM), (B) γ-tocotrienol (50 μM), or (C) both agents for 48 h. Whole-cell extracts were prepared and were analyzed by western blot analysis using antibodies against survivin, cIAP-1, cIAP-2, XIAP, Bcl-2 and β-actin as indicated. Although docetaxel stimulated the expression of these anti-apoptotic proteins, γ-tocotrienol slightly suppressed the expression of these proteins. On the other hand, simultaneous treatment of cells with both agents significantly inhibited the expression of anti-apoptotic proteins. As a loading control for the protein samples, the expression of β-actin is demonstrated. Results are representative of 3 independent experiments.

    Journal: International Journal of Oncology

    Article Title: ?-tocotrienol enhances the chemosensitivity of human oral cancer cells to docetaxel through the downregulation of the expression of NF-?B-regulated anti-apoptotic gene products

    doi: 10.3892/ijo.2012.1692

    Figure Lengend Snippet: γ-tocotrienol inhibition of the anti-apoptotic gene products, survivin, cIAP-1, cIAP-2, XIAP and Bcl-2. Cells were left untreated or incubated with either (A) docetaxel (0.5 nM), (B) γ-tocotrienol (50 μM), or (C) both agents for 48 h. Whole-cell extracts were prepared and were analyzed by western blot analysis using antibodies against survivin, cIAP-1, cIAP-2, XIAP, Bcl-2 and β-actin as indicated. Although docetaxel stimulated the expression of these anti-apoptotic proteins, γ-tocotrienol slightly suppressed the expression of these proteins. On the other hand, simultaneous treatment of cells with both agents significantly inhibited the expression of anti-apoptotic proteins. As a loading control for the protein samples, the expression of β-actin is demonstrated. Results are representative of 3 independent experiments.

    Article Snippet: Cells (5×103 cells per well) were grown on 96-well plates (Falcon Labware, Lincoln Park, NJ) in DMEM supplemented with 10% serum in the presence or absence of docetaxel (0.5, 1, and 2 nM) (obtained from Sigma, St. Louis, MO) and γ-tocotrienol (50, 75, and 100 μM) (obtained from Eizai Food & Chemical Co., Tokyo, Japan, with a purity exceeding 98.7%) alone, or both for 6 days.

    Techniques: Inhibition, Incubation, Western Blot, Expressing

    Akt is the regulator of the synergism, although resveratrol downregulates docetaxel-induced upregulation of Akt and MAPK pathways in SK-BR-3 cells. ( a ) Kinetics of docetaxel-induced activation of Akt. Cells were treated with docetaxel for different time intervals (0–2 h). The whole-cell lysate was immunoblotted against phospho-Akt (ser473) antibody. ( b ) Resveratrol-mediated downregulation of docetaxel-induced activation of Akt. Western blot analyses were performed with anti-phospho-Akt (ser473) using whole-cell lysates prepared after 30 min exposure to docetaxel. ( c ) Effect of resveratrol on docetaxel-induced upregulation of phospho-Bad. Western blot analysis was performed against anti-phospho-Bad (ser136). ( d ) Kinetics of activation of MAPKs by docetaxel (0–2 h). The whole-cell lysate was immunoblotted against phospho-specific antibodies of ERK1/2, JNK and p38. ( e ) Resveratrol downregulates docetaxel-induced upregulation of various MAPKs. β -Actin was used as loading control in all cases. ( f ) Inhibition of docetaxel-induced activation of AP-1 by resveratrol. Nuclear extracts prepared after exposing the cells to docetaxel and resveratrol, either alone or in combination for a period of 1 h, were assayed for AP-1 activation by EMSA. ( g ) Super-shift analysis using anti-c-jun antibody to indicate band specificity. ( h ) Kinetics of docetaxel-induced activation of NF- κ B. Nuclear extracts were prepared after exposing the cells to 1 nM docetaxel for different time intervals (0–3 h) and NF- κ B status was assessed by EMSA. ( i ) Individual and combined effects of docetaxel and resveratrol for a period of 30 min on NF- κ B activation. NF- κ B activation was assayed by EMSA. ( j ) Effect of docetaxel and resveratrol, alone or in combination, in cells treated with Akt and MAPKs inhibitors. Cells (5×10 3 ) in triplicates were pretreated with resveratrol, LY294002 (1 μ M), U0126 (5 μ M), SP600125 (5 μ M) and SB203580 (1 μ M), followed by docetaxel treatment for 48 h and subjected to MTT assay. Inhibition status of Akt and various MAPKs were shown in inset.

    Journal: Cell Death Discovery

    Article Title: Resveratrol chemosensitizes HER-2-overexpressing breast cancer cells to docetaxel chemoresistance by inhibiting docetaxel-mediated activation of HER-2–Akt axis

    doi: 10.1038/cddiscovery.2015.61

    Figure Lengend Snippet: Akt is the regulator of the synergism, although resveratrol downregulates docetaxel-induced upregulation of Akt and MAPK pathways in SK-BR-3 cells. ( a ) Kinetics of docetaxel-induced activation of Akt. Cells were treated with docetaxel for different time intervals (0–2 h). The whole-cell lysate was immunoblotted against phospho-Akt (ser473) antibody. ( b ) Resveratrol-mediated downregulation of docetaxel-induced activation of Akt. Western blot analyses were performed with anti-phospho-Akt (ser473) using whole-cell lysates prepared after 30 min exposure to docetaxel. ( c ) Effect of resveratrol on docetaxel-induced upregulation of phospho-Bad. Western blot analysis was performed against anti-phospho-Bad (ser136). ( d ) Kinetics of activation of MAPKs by docetaxel (0–2 h). The whole-cell lysate was immunoblotted against phospho-specific antibodies of ERK1/2, JNK and p38. ( e ) Resveratrol downregulates docetaxel-induced upregulation of various MAPKs. β -Actin was used as loading control in all cases. ( f ) Inhibition of docetaxel-induced activation of AP-1 by resveratrol. Nuclear extracts prepared after exposing the cells to docetaxel and resveratrol, either alone or in combination for a period of 1 h, were assayed for AP-1 activation by EMSA. ( g ) Super-shift analysis using anti-c-jun antibody to indicate band specificity. ( h ) Kinetics of docetaxel-induced activation of NF- κ B. Nuclear extracts were prepared after exposing the cells to 1 nM docetaxel for different time intervals (0–3 h) and NF- κ B status was assessed by EMSA. ( i ) Individual and combined effects of docetaxel and resveratrol for a period of 30 min on NF- κ B activation. NF- κ B activation was assayed by EMSA. ( j ) Effect of docetaxel and resveratrol, alone or in combination, in cells treated with Akt and MAPKs inhibitors. Cells (5×10 3 ) in triplicates were pretreated with resveratrol, LY294002 (1 μ M), U0126 (5 μ M), SP600125 (5 μ M) and SB203580 (1 μ M), followed by docetaxel treatment for 48 h and subjected to MTT assay. Inhibition status of Akt and various MAPKs were shown in inset.

    Article Snippet: All other chemicals including docetaxel were purchased from Sigma Chemicals (St. Louis, MO, USA).

    Techniques: Activation Assay, Western Blot, Inhibition, MTT Assay

    Docetaxel-induced upregulation of XIAP, survivin and Bcl-2 are downregulated by resveratrol in SK-BR-3 cells. ( a ) Effect of HER-2 inhibition on activation of Akt and downstream target survivin. ( b ) Effect of HER-2 inhibition on docetaxel-induced upregulation of phospho-Akt. ( c ) Effect of Akt inhibition on docetaxel-induced activation of HER-2. Western blotting analysis was done using specific antibodies against each molecule and β -actin was used as loading control in all cases. ( d ) Resveratrol-mediated inhibition of kinase activity of all three classes of Akt. The study was performed using z-lite biochemical assay platform. ( e ) Kinetics of docetaxel-induced activation of survivin and the inhibition of the same by synergistic combination of resveratrol and docetaxel. The cells were treated with docetaxel for different time intervals (0–24 h) and the whole-cell lysates were immunoblotted against anti-survivin. The cells were pretreated with resveratrol for 24 h followed by docetaxel alone or in combination with resveratrol for 8 h and immunoblotted against survivin antibody. ( f ) Kinetics of docetaxel-induced activation of XIAP and inhibition of the same by synergistic combination of resveratrol and docetaxel. The cells were treated with docetaxel for different time intervals (0–24 h). Western blotting analysis was done against anti-XIAP. The cells were pretreated with resveratrol for 24 h followed by docetaxel alone or in combination with resveratrol for another 16 h and western blot analysis was performed using antibody against anti-XIAP. ( g ) Kinetics of docetaxel-induced activation of Bcl-2 and inhibition of the same by synergistic combination of resveratrol and docetaxel. Cells were treated with of docetaxel for different time intervals (0–48 h). Western blot analysis was performed against anti-Bcl-2. Cells were pretreated with resveratrol followed by combination of resveratrol and docetaxel for 4 h. Whole-cell lysate was immunoblotted using anti-Bcl-2. β -Actin was used as loading control in all the experiments.

    Journal: Cell Death Discovery

    Article Title: Resveratrol chemosensitizes HER-2-overexpressing breast cancer cells to docetaxel chemoresistance by inhibiting docetaxel-mediated activation of HER-2–Akt axis

    doi: 10.1038/cddiscovery.2015.61

    Figure Lengend Snippet: Docetaxel-induced upregulation of XIAP, survivin and Bcl-2 are downregulated by resveratrol in SK-BR-3 cells. ( a ) Effect of HER-2 inhibition on activation of Akt and downstream target survivin. ( b ) Effect of HER-2 inhibition on docetaxel-induced upregulation of phospho-Akt. ( c ) Effect of Akt inhibition on docetaxel-induced activation of HER-2. Western blotting analysis was done using specific antibodies against each molecule and β -actin was used as loading control in all cases. ( d ) Resveratrol-mediated inhibition of kinase activity of all three classes of Akt. The study was performed using z-lite biochemical assay platform. ( e ) Kinetics of docetaxel-induced activation of survivin and the inhibition of the same by synergistic combination of resveratrol and docetaxel. The cells were treated with docetaxel for different time intervals (0–24 h) and the whole-cell lysates were immunoblotted against anti-survivin. The cells were pretreated with resveratrol for 24 h followed by docetaxel alone or in combination with resveratrol for 8 h and immunoblotted against survivin antibody. ( f ) Kinetics of docetaxel-induced activation of XIAP and inhibition of the same by synergistic combination of resveratrol and docetaxel. The cells were treated with docetaxel for different time intervals (0–24 h). Western blotting analysis was done against anti-XIAP. The cells were pretreated with resveratrol for 24 h followed by docetaxel alone or in combination with resveratrol for another 16 h and western blot analysis was performed using antibody against anti-XIAP. ( g ) Kinetics of docetaxel-induced activation of Bcl-2 and inhibition of the same by synergistic combination of resveratrol and docetaxel. Cells were treated with of docetaxel for different time intervals (0–48 h). Western blot analysis was performed against anti-Bcl-2. Cells were pretreated with resveratrol followed by combination of resveratrol and docetaxel for 4 h. Whole-cell lysate was immunoblotted using anti-Bcl-2. β -Actin was used as loading control in all the experiments.

    Article Snippet: All other chemicals including docetaxel were purchased from Sigma Chemicals (St. Louis, MO, USA).

    Techniques: Inhibition, Activation Assay, Western Blot, Activity Assay

    Resveratrol enhances docetaxel-induced apoptosis in SK-BR-3 cells. ( a ) Cells were treated with resveratrol and/or docetaxel for 16 h and stained for Annexin V–propidium iodide (PI) positivity. Annexin V-positive cells in different fields were counted and the average was taken. The green-stained cells are those that have taken only the Annexin V–FITC stain and represent initial stages of apoptosis, and the red-stained cells are those that have taken up both Annexin–FITC and PI, which indicates nuclear membrane damage, and hence represent later stages of apoptosis. Representative histograms indicate percentage of annexin-positive cells. *** P -value ≤0.001.( b – e ) Resveratrol-mediated enhancement of docetaxel-induced caspase activation. Whole cell lysate of cells treated with docetaxel and/or resveratrol for 48 h were blotted against caspase antibodies. ( f ) Resveratrol enhances docetaxel-induced PARP cleavage. Whole-cell extracts were blotted against anti-PARP antibody. ( g ) Effect of docetaxel and resveratrol, alone or in combination, on cell cycle. Cells were collected 48 h post drug treatment, fixed in alcohol, stained with propidium iodide and assayed for DNA content by flow cytometry. ( h ) The effect of docetaxel and/or resveratrol on inter-nucleosomal DNA fragmentation. Cells were treated with docetaxel and/or resveratrol for 48 h, DNA was isolated, run on an agarose gel and visualized. All experiments were repeated at least three times to confirm the reproducibility.

    Journal: Cell Death Discovery

    Article Title: Resveratrol chemosensitizes HER-2-overexpressing breast cancer cells to docetaxel chemoresistance by inhibiting docetaxel-mediated activation of HER-2–Akt axis

    doi: 10.1038/cddiscovery.2015.61

    Figure Lengend Snippet: Resveratrol enhances docetaxel-induced apoptosis in SK-BR-3 cells. ( a ) Cells were treated with resveratrol and/or docetaxel for 16 h and stained for Annexin V–propidium iodide (PI) positivity. Annexin V-positive cells in different fields were counted and the average was taken. The green-stained cells are those that have taken only the Annexin V–FITC stain and represent initial stages of apoptosis, and the red-stained cells are those that have taken up both Annexin–FITC and PI, which indicates nuclear membrane damage, and hence represent later stages of apoptosis. Representative histograms indicate percentage of annexin-positive cells. *** P -value ≤0.001.( b – e ) Resveratrol-mediated enhancement of docetaxel-induced caspase activation. Whole cell lysate of cells treated with docetaxel and/or resveratrol for 48 h were blotted against caspase antibodies. ( f ) Resveratrol enhances docetaxel-induced PARP cleavage. Whole-cell extracts were blotted against anti-PARP antibody. ( g ) Effect of docetaxel and resveratrol, alone or in combination, on cell cycle. Cells were collected 48 h post drug treatment, fixed in alcohol, stained with propidium iodide and assayed for DNA content by flow cytometry. ( h ) The effect of docetaxel and/or resveratrol on inter-nucleosomal DNA fragmentation. Cells were treated with docetaxel and/or resveratrol for 48 h, DNA was isolated, run on an agarose gel and visualized. All experiments were repeated at least three times to confirm the reproducibility.

    Article Snippet: All other chemicals including docetaxel were purchased from Sigma Chemicals (St. Louis, MO, USA).

    Techniques: Staining, Activation Assay, Flow Cytometry, Cytometry, Isolation, Agarose Gel Electrophoresis

    Resveratrol downregulates docetaxel-induced activation of HER-2. ( a Kinetics of docetaxel (1 nM)-induced activation of HER-2 in SK-BR-3 cells (0–48 h). The whole-cell lysate of docetaxel-treated cells at different time intervals was immunoblotted against HER-2 antibody. ( b and c ) Effect of resveratrol treatment on docetaxel-induced overexpression and activation of HER-2 in SK-BR-3 cells. Cells pretreated with resveratrol for 24 h were further exposed to docetaxel and/or resveratrol for another 24 h and immunoblotted against specific antibodies. β -Tubulin is used as loading control. ( d ) Effect of HER-2 inhibition on synergism in SK-BR-3 cells. Cells were transiently transfected with vector control and DN-HER-2, respectively. ( e ) Effect of ectopic expression of HER-2 on synergism in MDA-MB-231 cells. Cells were transfected with vector control and WT-HER-2, respectively. The efficacy of transfection was confirmed using western blotting and cell viability was assessed in the transfected cells using MTT assay after treatment with docetaxel and resveratrol alone, or in combination for 48 h in both cases.

    Journal: Cell Death Discovery

    Article Title: Resveratrol chemosensitizes HER-2-overexpressing breast cancer cells to docetaxel chemoresistance by inhibiting docetaxel-mediated activation of HER-2–Akt axis

    doi: 10.1038/cddiscovery.2015.61

    Figure Lengend Snippet: Resveratrol downregulates docetaxel-induced activation of HER-2. ( a Kinetics of docetaxel (1 nM)-induced activation of HER-2 in SK-BR-3 cells (0–48 h). The whole-cell lysate of docetaxel-treated cells at different time intervals was immunoblotted against HER-2 antibody. ( b and c ) Effect of resveratrol treatment on docetaxel-induced overexpression and activation of HER-2 in SK-BR-3 cells. Cells pretreated with resveratrol for 24 h were further exposed to docetaxel and/or resveratrol for another 24 h and immunoblotted against specific antibodies. β -Tubulin is used as loading control. ( d ) Effect of HER-2 inhibition on synergism in SK-BR-3 cells. Cells were transiently transfected with vector control and DN-HER-2, respectively. ( e ) Effect of ectopic expression of HER-2 on synergism in MDA-MB-231 cells. Cells were transfected with vector control and WT-HER-2, respectively. The efficacy of transfection was confirmed using western blotting and cell viability was assessed in the transfected cells using MTT assay after treatment with docetaxel and resveratrol alone, or in combination for 48 h in both cases.

    Article Snippet: All other chemicals including docetaxel were purchased from Sigma Chemicals (St. Louis, MO, USA).

    Techniques: Activation Assay, Over Expression, Inhibition, Transfection, Plasmid Preparation, Dominant Negative Mutation, Expressing, Multiple Displacement Amplification, Western Blot, MTT Assay

    Resveratrol sensitizes breast cancer cells to docetaxel-induced cytotoxicity, while being non-toxic to normal immortalized breast cells. ( a ) Dose-dependent cytotoxicity of docetaxel (0.1–10 nM) on breast cancer cells with varied receptor status. ( b ) Effect of different concentrations of resveratrol (10–25 μ M) on different breast cancer cells. ( c ) Effect of docetaxel (1 nM) and resveratrol (15 μ M), alone or in combination on different breast cancer cells. Cells (5×10 3 ) in triplicates were exposed to the indicated concentrations of the docetaxel for 48 h and subjected to MTT assay. Relative cell viability was determined as percentage absorbance over untreated control. ( d ) Effect of docetaxel and resveratrol, alone or in combination, on SK-BR-3 cells using [ 3 H] thymidine incorporation assay. Cells (5×10 3 ) in triplicates were exposed to the indicated concentrations of the drugs for 24 h and subjected to [ 3 H] thymidine incorporation assay. Relative cell viability was determined as percentage thymidine incorporation over control. ( e ) The combination is non-toxic to MCF10A as assessed by [ 3 H] thymidine incorporation assay. ( f ) Inhibitory effect of docetaxel and resveratrol, alone or in combination, on the clonogenic ability of SK-BR-3 cells (scale bar, 1 μ m/px).

    Journal: Cell Death Discovery

    Article Title: Resveratrol chemosensitizes HER-2-overexpressing breast cancer cells to docetaxel chemoresistance by inhibiting docetaxel-mediated activation of HER-2–Akt axis

    doi: 10.1038/cddiscovery.2015.61

    Figure Lengend Snippet: Resveratrol sensitizes breast cancer cells to docetaxel-induced cytotoxicity, while being non-toxic to normal immortalized breast cells. ( a ) Dose-dependent cytotoxicity of docetaxel (0.1–10 nM) on breast cancer cells with varied receptor status. ( b ) Effect of different concentrations of resveratrol (10–25 μ M) on different breast cancer cells. ( c ) Effect of docetaxel (1 nM) and resveratrol (15 μ M), alone or in combination on different breast cancer cells. Cells (5×10 3 ) in triplicates were exposed to the indicated concentrations of the docetaxel for 48 h and subjected to MTT assay. Relative cell viability was determined as percentage absorbance over untreated control. ( d ) Effect of docetaxel and resveratrol, alone or in combination, on SK-BR-3 cells using [ 3 H] thymidine incorporation assay. Cells (5×10 3 ) in triplicates were exposed to the indicated concentrations of the drugs for 24 h and subjected to [ 3 H] thymidine incorporation assay. Relative cell viability was determined as percentage thymidine incorporation over control. ( e ) The combination is non-toxic to MCF10A as assessed by [ 3 H] thymidine incorporation assay. ( f ) Inhibitory effect of docetaxel and resveratrol, alone or in combination, on the clonogenic ability of SK-BR-3 cells (scale bar, 1 μ m/px).

    Article Snippet: All other chemicals including docetaxel were purchased from Sigma Chemicals (St. Louis, MO, USA).

    Techniques: MTT Assay, Thymidine Incorporation Assay

    Proposed model for the synergistic effect of docetaxel and resveratrol. Resveratrol downregulates docetaxel-induced upregulation of HER-2, Akt and MAPKs, while that of NF- κ B is unaffected. The study postulates that docetaxel-induced upregulation of HER-2–Akt signaling and the downregulation of the same by resveratrol is the main mechanism governing the synergistic effect of docetaxel and resveratrol in HER-2-overexpressing breast cancer cells. Although MAPK pathway does not regulate the synergism, it is getting activated by docetaxel and downregulated by resveratrol. The bold lines indicate the signaling pathways regulating the synergism, whereas the dotted lines represent those that do not have any role in the same.

    Journal: Cell Death Discovery

    Article Title: Resveratrol chemosensitizes HER-2-overexpressing breast cancer cells to docetaxel chemoresistance by inhibiting docetaxel-mediated activation of HER-2–Akt axis

    doi: 10.1038/cddiscovery.2015.61

    Figure Lengend Snippet: Proposed model for the synergistic effect of docetaxel and resveratrol. Resveratrol downregulates docetaxel-induced upregulation of HER-2, Akt and MAPKs, while that of NF- κ B is unaffected. The study postulates that docetaxel-induced upregulation of HER-2–Akt signaling and the downregulation of the same by resveratrol is the main mechanism governing the synergistic effect of docetaxel and resveratrol in HER-2-overexpressing breast cancer cells. Although MAPK pathway does not regulate the synergism, it is getting activated by docetaxel and downregulated by resveratrol. The bold lines indicate the signaling pathways regulating the synergism, whereas the dotted lines represent those that do not have any role in the same.

    Article Snippet: All other chemicals including docetaxel were purchased from Sigma Chemicals (St. Louis, MO, USA).

    Techniques:

    CXCR4si increased the sensitivity of VCaP and PC-3 cells to docetaxel and reduced cell migration trend when combined with docetaxel. a . Western blot in VCaP and PC-3 cells to evaluate the efficacy of CXCR4si7 and CXCR4si8 siRNAs. The results are presented as the mean ± standard error from three independent experiments. Quantitative data of western blots on 2 different CXCR4siRNAs for validation and determination of their effect size are provided in Supplementary Data. b . CXCR4 knockdown decreased the survival of VCaP cells when was combined with GLIPR1-ΔTM ( p

    Journal: Molecular Cancer

    Article Title: GLIPR1-ΔTM synergizes with docetaxel in cell death and suppresses resistance to docetaxel in prostate cancer cells

    doi: 10.1186/s12943-015-0395-0

    Figure Lengend Snippet: CXCR4si increased the sensitivity of VCaP and PC-3 cells to docetaxel and reduced cell migration trend when combined with docetaxel. a . Western blot in VCaP and PC-3 cells to evaluate the efficacy of CXCR4si7 and CXCR4si8 siRNAs. The results are presented as the mean ± standard error from three independent experiments. Quantitative data of western blots on 2 different CXCR4siRNAs for validation and determination of their effect size are provided in Supplementary Data. b . CXCR4 knockdown decreased the survival of VCaP cells when was combined with GLIPR1-ΔTM ( p

    Article Snippet: Docetaxel and AMD3100, a CXCR4 inhibitor, were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Migration, Western Blot

    Docetaxel GLIPR1-ΔΤΜ combination treatment inhibited growth and metastasis in VCaP orthotopic xenografts. a . Nude mice bearing VCaP xenografts were treated with docetaxel, GLIPR1-ΔΤΜ, or both. Combination treatment significantly decreased the IVIS signal (photons/s) than GLIPR1-ΔΤΜ alone did ( p = 0.012) but docetaxel alone did not ( p = 0.16). b . Combination treatment decreased significantly the wet weight of VCaP xenografts rather thad GLIPR1-ΔΤΜ alone ( p = 0.0025) and docetaxel alone ( p = 0.028) did. c . Mouse lymph node tissues were stained for cytokeratin 18 to evaluate the presence of metastatic VCaP cells. The combination of docetaxel and GLIPR1-ΔΤΜ resulted in a lower incidence of LN metastasis than control treatment did ( p = 0.04), whereas single-agent treatments did not result in a significant reduction

    Journal: Molecular Cancer

    Article Title: GLIPR1-ΔTM synergizes with docetaxel in cell death and suppresses resistance to docetaxel in prostate cancer cells

    doi: 10.1186/s12943-015-0395-0

    Figure Lengend Snippet: Docetaxel GLIPR1-ΔΤΜ combination treatment inhibited growth and metastasis in VCaP orthotopic xenografts. a . Nude mice bearing VCaP xenografts were treated with docetaxel, GLIPR1-ΔΤΜ, or both. Combination treatment significantly decreased the IVIS signal (photons/s) than GLIPR1-ΔΤΜ alone did ( p = 0.012) but docetaxel alone did not ( p = 0.16). b . Combination treatment decreased significantly the wet weight of VCaP xenografts rather thad GLIPR1-ΔΤΜ alone ( p = 0.0025) and docetaxel alone ( p = 0.028) did. c . Mouse lymph node tissues were stained for cytokeratin 18 to evaluate the presence of metastatic VCaP cells. The combination of docetaxel and GLIPR1-ΔΤΜ resulted in a lower incidence of LN metastasis than control treatment did ( p = 0.04), whereas single-agent treatments did not result in a significant reduction

    Article Snippet: Docetaxel and AMD3100, a CXCR4 inhibitor, were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Mouse Assay, Staining

    GLIPR1-ΔTM reduced VCaP and PC-3 cells’ survival synergistically with docetaxel. a . Docetaxel dose–response curves of VCaP, PC-3, and RWPE-1 cells treated for 48 h, determined using MTS assay. At 0.5nM, docetaxel significantly decreased cell survival in the three cell lines. The IC50 was 69.8nM for VCaP cells and 70.5nM for PC-3 cells. b . GLIPR1-ΔTM dose–response curves of VCaP, PC-3, and RWPE-1 cells treated for 48 h, determined using MTS assay. At 2.5 μg/ml and 10 μg/ml, GLIPR1-ΔTM significantly decreased survival of VCaP and PC-3 cells, respectively. Only 160 μg/ml decreased survival of RWPE-1 cells. The IC50 was 34.8 μg/ml for VCaP cells and 154 μg/ml for PC-3 cells. c . The addition of 10 μg/ml GLIPR1-ΔTM increased the efficacies of all doses of docetaxel in inhibiting VCaP cells’ survival, based on MTS assay. d . The addition of 10 μg/ml GLIPR1-ΔTM increased the efficacies of 0.5, 1, and 2nM docetaxel in inhibiting PC-3 cells’ survival as determined by MTS assay, and the addition of 80 μg/ml GLIPR1-ΔTM increased the efficacies of 10, 20, and 50nM docetaxel. e . Isobologram analysis showed that GLIPR1-ΔTM and docetaxel synergistically induced cell death in VCaP cells. f . Isobologram analysis showed that GLIPR1-ΔTM and docetaxel induced cell death synergistically in PC-3 cells. The results are presented as the mean ± standard error from at least three independent experiments

    Journal: Molecular Cancer

    Article Title: GLIPR1-ΔTM synergizes with docetaxel in cell death and suppresses resistance to docetaxel in prostate cancer cells

    doi: 10.1186/s12943-015-0395-0

    Figure Lengend Snippet: GLIPR1-ΔTM reduced VCaP and PC-3 cells’ survival synergistically with docetaxel. a . Docetaxel dose–response curves of VCaP, PC-3, and RWPE-1 cells treated for 48 h, determined using MTS assay. At 0.5nM, docetaxel significantly decreased cell survival in the three cell lines. The IC50 was 69.8nM for VCaP cells and 70.5nM for PC-3 cells. b . GLIPR1-ΔTM dose–response curves of VCaP, PC-3, and RWPE-1 cells treated for 48 h, determined using MTS assay. At 2.5 μg/ml and 10 μg/ml, GLIPR1-ΔTM significantly decreased survival of VCaP and PC-3 cells, respectively. Only 160 μg/ml decreased survival of RWPE-1 cells. The IC50 was 34.8 μg/ml for VCaP cells and 154 μg/ml for PC-3 cells. c . The addition of 10 μg/ml GLIPR1-ΔTM increased the efficacies of all doses of docetaxel in inhibiting VCaP cells’ survival, based on MTS assay. d . The addition of 10 μg/ml GLIPR1-ΔTM increased the efficacies of 0.5, 1, and 2nM docetaxel in inhibiting PC-3 cells’ survival as determined by MTS assay, and the addition of 80 μg/ml GLIPR1-ΔTM increased the efficacies of 10, 20, and 50nM docetaxel. e . Isobologram analysis showed that GLIPR1-ΔTM and docetaxel synergistically induced cell death in VCaP cells. f . Isobologram analysis showed that GLIPR1-ΔTM and docetaxel induced cell death synergistically in PC-3 cells. The results are presented as the mean ± standard error from at least three independent experiments

    Article Snippet: Docetaxel and AMD3100, a CXCR4 inhibitor, were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: MTS Assay

    GLIPR1-ΔTM increased the apoptotic effect of docetaxel in VCaP and PC-3 cells. a . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel’s doses in VCaP cells, measured by DAPI staining. b . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel doses in PC-3 cells, measured by DAPI staining. c . GLIPR1-ΔΤΜ did not increase the apoptotic effect of any docetaxel dose in RWPE-1 cells, while 1nM docetaxel significantly increased the percentage of apoptotic cells, based on DAPI staining. d . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel doses in VCaP cells, measured by DNA fragmentation. e . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel doses in PC-3 cells, measured by DNA fragmentation. f . GLIPR1-ΔΤΜ did not increase the apoptotic effect of any docetaxel dose in RWPE-1 cells, while 0.5nM docetaxel significantly increased the percentage of apoptotic cells, based on DNA fragmentation. The results are presented as the mean ± standard error from at least three independent experiments

    Journal: Molecular Cancer

    Article Title: GLIPR1-ΔTM synergizes with docetaxel in cell death and suppresses resistance to docetaxel in prostate cancer cells

    doi: 10.1186/s12943-015-0395-0

    Figure Lengend Snippet: GLIPR1-ΔTM increased the apoptotic effect of docetaxel in VCaP and PC-3 cells. a . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel’s doses in VCaP cells, measured by DAPI staining. b . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel doses in PC-3 cells, measured by DAPI staining. c . GLIPR1-ΔΤΜ did not increase the apoptotic effect of any docetaxel dose in RWPE-1 cells, while 1nM docetaxel significantly increased the percentage of apoptotic cells, based on DAPI staining. d . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel doses in VCaP cells, measured by DNA fragmentation. e . The addition of 10 μg/ml GLIPR1-ΔΤΜ increased the apoptotic effects of all docetaxel doses in PC-3 cells, measured by DNA fragmentation. f . GLIPR1-ΔΤΜ did not increase the apoptotic effect of any docetaxel dose in RWPE-1 cells, while 0.5nM docetaxel significantly increased the percentage of apoptotic cells, based on DNA fragmentation. The results are presented as the mean ± standard error from at least three independent experiments

    Article Snippet: Docetaxel and AMD3100, a CXCR4 inhibitor, were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Staining

    GLIPR1-ΔΤΜ synergized with docetaxel in activating JNK/c-Jun through JNK phosphorylation, whereas addition of GLIPR1-ΔΤΜ to docetaxel reduced ERK1/2 signaling. a , b . VCaP and PC-3 cells were treated with 1nM docetaxel, 10 μg/ml GLIPR1-ΔTM, or both and then, we added JNK inhibitor (SP600125) 1 μΜ to single agents or to their combination. Total treatment administration lasted for 24 h (VCaP cells) or 48 h (PC-3 cells), and the effects on JNK and ERK1/2 signaling were evaluated via western blot. JNK phosphorylation was increased synergistically with the combination of docetaxel and GLIPR1-ΔΤΜ, a pathway that leads to apoptosis; whereas ERK1/2 phosphorylation, which results in drug resistance and migration through c-Myc-CXCR4, was reduced by administration of this combination. These activities were reversed by the JNK inhibitor, SP600125. Western blot experiments were conducted three times and the quantitative data are presented as supplementary data. c , d . Under the same conditions and treatments in both cell lines, we performed MTS and DNA fragmentation assay to evaluate the percentage of survived cells and cell apoptosis, respectively. We found that, in both cell lines, the combination treatment of docetaxel and GLIPR1-ΔΤΜ resulted in statistically significant decrease in the percentage of survived cells and increase of apoptotic cells (VCaP: p

    Journal: Molecular Cancer

    Article Title: GLIPR1-ΔTM synergizes with docetaxel in cell death and suppresses resistance to docetaxel in prostate cancer cells

    doi: 10.1186/s12943-015-0395-0

    Figure Lengend Snippet: GLIPR1-ΔΤΜ synergized with docetaxel in activating JNK/c-Jun through JNK phosphorylation, whereas addition of GLIPR1-ΔΤΜ to docetaxel reduced ERK1/2 signaling. a , b . VCaP and PC-3 cells were treated with 1nM docetaxel, 10 μg/ml GLIPR1-ΔTM, or both and then, we added JNK inhibitor (SP600125) 1 μΜ to single agents or to their combination. Total treatment administration lasted for 24 h (VCaP cells) or 48 h (PC-3 cells), and the effects on JNK and ERK1/2 signaling were evaluated via western blot. JNK phosphorylation was increased synergistically with the combination of docetaxel and GLIPR1-ΔΤΜ, a pathway that leads to apoptosis; whereas ERK1/2 phosphorylation, which results in drug resistance and migration through c-Myc-CXCR4, was reduced by administration of this combination. These activities were reversed by the JNK inhibitor, SP600125. Western blot experiments were conducted three times and the quantitative data are presented as supplementary data. c , d . Under the same conditions and treatments in both cell lines, we performed MTS and DNA fragmentation assay to evaluate the percentage of survived cells and cell apoptosis, respectively. We found that, in both cell lines, the combination treatment of docetaxel and GLIPR1-ΔΤΜ resulted in statistically significant decrease in the percentage of survived cells and increase of apoptotic cells (VCaP: p

    Article Snippet: Docetaxel and AMD3100, a CXCR4 inhibitor, were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Western Blot, Migration, DNA Fragmentation Assay

    Time-dependent curves of docetaxel and GLIPR1-ΔTM efficacies in VCaP and PC-3 cells treated with 1, 2, or 5nM docetaxel or 10, 20, or 40 μg/ml GLIPR1-ΔTM. a , b . 1nM docetaxel and 10 μg/ml GLIPR1-ΔΤΜ significantly decreased survival of VCaP cells at 24 h ( p = 0.03 for docetaxel and p = 0.001 for GLIPR1-ΔTM). c , d . Time-dependent curves of docetaxel and GLIPR1-ΔTM efficacies in PC-3 cells treated with 1, 2, or 5nM docetaxel or 10, 20, or 40 μg/ml GLIPR1-ΔTM showed that 1nM docetaxel and 10 μg/ml GLIPR1-ΔΤΜ significantly decreased survival of these cells at 48 h ( p

    Journal: Molecular Cancer

    Article Title: GLIPR1-ΔTM synergizes with docetaxel in cell death and suppresses resistance to docetaxel in prostate cancer cells

    doi: 10.1186/s12943-015-0395-0

    Figure Lengend Snippet: Time-dependent curves of docetaxel and GLIPR1-ΔTM efficacies in VCaP and PC-3 cells treated with 1, 2, or 5nM docetaxel or 10, 20, or 40 μg/ml GLIPR1-ΔTM. a , b . 1nM docetaxel and 10 μg/ml GLIPR1-ΔΤΜ significantly decreased survival of VCaP cells at 24 h ( p = 0.03 for docetaxel and p = 0.001 for GLIPR1-ΔTM). c , d . Time-dependent curves of docetaxel and GLIPR1-ΔTM efficacies in PC-3 cells treated with 1, 2, or 5nM docetaxel or 10, 20, or 40 μg/ml GLIPR1-ΔTM showed that 1nM docetaxel and 10 μg/ml GLIPR1-ΔΤΜ significantly decreased survival of these cells at 48 h ( p

    Article Snippet: Docetaxel and AMD3100, a CXCR4 inhibitor, were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques:

    Effects of docetaxel on ( A ) SKOV3 and SKOV3-TR or ( B ) HeyA8 and HeyA8-MDR ovarian cancer cells. The percentage of apoptosis was determined by terminal deoxynucleotidyl transferase – mediated nick end labeling. Cells were treated with or without

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effects of docetaxel on ( A ) SKOV3 and SKOV3-TR or ( B ) HeyA8 and HeyA8-MDR ovarian cancer cells. The percentage of apoptosis was determined by terminal deoxynucleotidyl transferase – mediated nick end labeling. Cells were treated with or without

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques: End Labeling

    Effect of docetaxel on ovarian cancer cell growth: ( A ) HeyA8 and HeyA8-MDR or ( B ) SKOV3 or SKOV3-TR cells were plated in 96-well plates and subsequently incubated with increasing concentrations of docetaxel for 96 hours, and cell viability was determined.

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effect of docetaxel on ovarian cancer cell growth: ( A ) HeyA8 and HeyA8-MDR or ( B ) SKOV3 or SKOV3-TR cells were plated in 96-well plates and subsequently incubated with increasing concentrations of docetaxel for 96 hours, and cell viability was determined.

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques: Incubation

    Effect of docetaxel on caspase-3 ( A ), caspase-8 ( B ), and caspase-9 ( C ) activity. Ovarian cancer cells were treated with docetaxel for 24 hours followed by fluorometric profiling of caspase activity using a commercially available kit. Columns, means of

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effect of docetaxel on caspase-3 ( A ), caspase-8 ( B ), and caspase-9 ( C ) activity. Ovarian cancer cells were treated with docetaxel for 24 hours followed by fluorometric profiling of caspase activity using a commercially available kit. Columns, means of

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques: Activity Assay

    Docetaxel treatment results in FAK cleavage. The taxane-sensitive ( A ) and taxane-resistant ( B ) SKOV3 cells were cultured in the presence of docetaxel for 48 or 72 hours. Western blot analysis for FAK in SKOV3 and SKOV3-TR cells in the presence or absence

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Docetaxel treatment results in FAK cleavage. The taxane-sensitive ( A ) and taxane-resistant ( B ) SKOV3 cells were cultured in the presence of docetaxel for 48 or 72 hours. Western blot analysis for FAK in SKOV3 and SKOV3-TR cells in the presence or absence

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques: Cell Culture, Western Blot

    Effect of FAK silencing on docetaxel-sensitivity in ovarian cancer cell lines. A, Western blot analysis of SKOV3 whole-cell lysates probed with anti-FAK and anti-actin monoclonal antibodies. Bottom, densitometry results. Control siRNA did not significantly

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effect of FAK silencing on docetaxel-sensitivity in ovarian cancer cell lines. A, Western blot analysis of SKOV3 whole-cell lysates probed with anti-FAK and anti-actin monoclonal antibodies. Bottom, densitometry results. Control siRNA did not significantly

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques: Western Blot

    Docetaxel-mediated apoptosis with or without FAK siRNA in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR ovarian cancer cells. Columns, means of three independent experiments; bars, SE. Doc, docetaxel.

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Docetaxel-mediated apoptosis with or without FAK siRNA in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR ovarian cancer cells. Columns, means of three independent experiments; bars, SE. Doc, docetaxel.

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques:

    FAK gene silencing potentiates docetaxel-induced caspase-3 activity in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR cells. Columns, means of three experiments; bars, SE. *, P

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: FAK gene silencing potentiates docetaxel-induced caspase-3 activity in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR cells. Columns, means of three experiments; bars, SE. *, P

    Article Snippet: Docetaxel was purchased from Sanofi-Aventis (Bridgewater, NJ).

    Techniques: Activity Assay

    Ki67 and TUNEL expression in tumors treated with docetaxel and PSK. Mice with established prostate tumors were treated with oral saline daily, oral PSK (300 mg/kg) daily, i.p. docetaxel injection (2x weekly) or a combination of PSK and docetaxel for a period of 10–12 days. Tumors were stained for the Ki67 proliferation marker and TUNEL expression. Shown are representative staining results and summary graphs showing the number of tumor cells positive for each marker (mean ± SEM) viewed at magnification ×40 for a total of 10 views in each of 6–8 tumors (hpf, per high powered field; H E, hematoxylin and eosin staining). Combining PSK with docetaxel induced a reduction in proliferating Ki67 + cells compared to docetaxel alone (p

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: Ki67 and TUNEL expression in tumors treated with docetaxel and PSK. Mice with established prostate tumors were treated with oral saline daily, oral PSK (300 mg/kg) daily, i.p. docetaxel injection (2x weekly) or a combination of PSK and docetaxel for a period of 10–12 days. Tumors were stained for the Ki67 proliferation marker and TUNEL expression. Shown are representative staining results and summary graphs showing the number of tumor cells positive for each marker (mean ± SEM) viewed at magnification ×40 for a total of 10 views in each of 6–8 tumors (hpf, per high powered field; H E, hematoxylin and eosin staining). Combining PSK with docetaxel induced a reduction in proliferating Ki67 + cells compared to docetaxel alone (p

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: TUNEL Assay, Expressing, Mouse Assay, Injection, Staining, Marker

    Effects of PSK and docetaxel treatments on mRNA expression of markers of antitumor immune response. Groups of mice (n=8 control; n=5 docetaxel; n= 6 PSK; n=4 PSK + docetaxel) bearing established TRAMP-C2 tumors (12 days) were treated with either oral saline, subtherapeutic dose of docetaxel (5 mg/kg twice weekly), oral gavage of PSK (300 mg/kg), or a combination of docetaxel and PSK. RNA was extracted from isolated tumors and real-time RT-PCR performed using primers and probes for several different cytokines and antitumor immune response markers. Shown are the effects of the different treatment groups on mRNA expression of IFN-γ (A), IL-2 (B), and TNF-α (C). The values shown are relative cytokine mRNA per 1,000 copies of β-actin. Mice treated with PSK, either alone (p=0.04) or with docetaxel (p=0.01) significantly induced IFN-γ mRNA expression in TRAMP-C2 tumors compared to saline control.

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: Effects of PSK and docetaxel treatments on mRNA expression of markers of antitumor immune response. Groups of mice (n=8 control; n=5 docetaxel; n= 6 PSK; n=4 PSK + docetaxel) bearing established TRAMP-C2 tumors (12 days) were treated with either oral saline, subtherapeutic dose of docetaxel (5 mg/kg twice weekly), oral gavage of PSK (300 mg/kg), or a combination of docetaxel and PSK. RNA was extracted from isolated tumors and real-time RT-PCR performed using primers and probes for several different cytokines and antitumor immune response markers. Shown are the effects of the different treatment groups on mRNA expression of IFN-γ (A), IL-2 (B), and TNF-α (C). The values shown are relative cytokine mRNA per 1,000 copies of β-actin. Mice treated with PSK, either alone (p=0.04) or with docetaxel (p=0.01) significantly induced IFN-γ mRNA expression in TRAMP-C2 tumors compared to saline control.

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: Expressing, Mouse Assay, Isolation, Quantitative RT-PCR

    PSK combined with docetaxel enhances tumor infiltrating T cells. Mice bearing orthotopic TRAMP-C2 tumors were treated with either: 1) saline; 2) PSK (300 mg/kg daily; 3) docetaxel (5 mg/kg); or 4) PSK and docetaxel. Mice were sacrificed on day 12 after treatment and tumors were harvested. (A) Fluorescent immunohistochemistry was performed for CD4 + and CD8 + T cells (examples of positive cells indicated by arrows). Populations of both CD4 + (B) and CD8 + (C) T cell subsets were significantly higher in mice treated with docetaxel and PSK compared to PSK or docetaxel alone (p

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: PSK combined with docetaxel enhances tumor infiltrating T cells. Mice bearing orthotopic TRAMP-C2 tumors were treated with either: 1) saline; 2) PSK (300 mg/kg daily; 3) docetaxel (5 mg/kg); or 4) PSK and docetaxel. Mice were sacrificed on day 12 after treatment and tumors were harvested. (A) Fluorescent immunohistochemistry was performed for CD4 + and CD8 + T cells (examples of positive cells indicated by arrows). Populations of both CD4 + (B) and CD8 + (C) T cell subsets were significantly higher in mice treated with docetaxel and PSK compared to PSK or docetaxel alone (p

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: Mouse Assay, Immunohistochemistry

    PSK in combination with docetaxel enhances NK cell activity of splenocytes from TRAMP-C2 tumor bearing mice. Splenocytes from mice treated with saline control or PSK and docetaxel alone and in combination were isolated and cultured in vitro with YAC-1 murine tumor target cells and assessed for NK cell killing of YAC-1 cells. Splenic cells from animals treated with PSK and docetaxel combination therapy had the highest level of NK cell activity at all effector:target ratios. NK cell activity induced by DTX/PSK was significantly enhanced at the 50:1 E:T ratio (p=0.045 using a linear mixed model to analyze data from 17 mice).

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: PSK in combination with docetaxel enhances NK cell activity of splenocytes from TRAMP-C2 tumor bearing mice. Splenocytes from mice treated with saline control or PSK and docetaxel alone and in combination were isolated and cultured in vitro with YAC-1 murine tumor target cells and assessed for NK cell killing of YAC-1 cells. Splenic cells from animals treated with PSK and docetaxel combination therapy had the highest level of NK cell activity at all effector:target ratios. NK cell activity induced by DTX/PSK was significantly enhanced at the 50:1 E:T ratio (p=0.045 using a linear mixed model to analyze data from 17 mice).

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: Activity Assay, Mouse Assay, Isolation, Cell Culture, In Vitro

    Oral administration of PSK plus docetaxel induces tumor regression. C57BL/6 mice with established tumors (50 mg) were treated with oral saline control daily, oral PSK (300 mg/kg) daily, i.p. injection of docetaxel (2x weekly) or a combination of PSK and docetaxel for a period of 10–12 days. Mean tumor weight ± standard error measurement (mean ± SEM) are shown for each treatment group of mice (n=7/group). PSK combined with docetaxel significantly suppressed tumor growth as compared to saline control, PSK or docetaxel treatments alone (P

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: Oral administration of PSK plus docetaxel induces tumor regression. C57BL/6 mice with established tumors (50 mg) were treated with oral saline control daily, oral PSK (300 mg/kg) daily, i.p. injection of docetaxel (2x weekly) or a combination of PSK and docetaxel for a period of 10–12 days. Mean tumor weight ± standard error measurement (mean ± SEM) are shown for each treatment group of mice (n=7/group). PSK combined with docetaxel significantly suppressed tumor growth as compared to saline control, PSK or docetaxel treatments alone (P

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: Mouse Assay, Injection

    Change in murine weights with combination therapy. Mice used in the same experiment as in Fig. 2 were weighed every other day for 12 days. Data are shown as mouse weight (mean ± sem) of each group (n=7 per group). Saline control or docetaxel treatments alone led to significant loss in murine weight by day 12, while PSK alone or PSK combined with docetaxel treatment maintained murine weight over this time period.

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: Change in murine weights with combination therapy. Mice used in the same experiment as in Fig. 2 were weighed every other day for 12 days. Data are shown as mouse weight (mean ± sem) of each group (n=7 per group). Saline control or docetaxel treatments alone led to significant loss in murine weight by day 12, while PSK alone or PSK combined with docetaxel treatment maintained murine weight over this time period.

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: Mouse Assay

    WBC in tumor bearing mice treated with PSK and docetaxel. Groups of mice (n=7) bearing established TRAMP-C2 tumors (12 days) were treated with either oral saline, subtherapeutic dose of docetaxel (5 mg/kg twice weekly), oral gavage of PSK (300 mg/kg), or a combination of docetaxel and PSK. WBC were assessed at the end of the experiment (mean ± SEM). Docetaxel alone induced a 25% reduction in WBC. Addition of PSK decreased the docetaxel-suppressive effect by ~50%, a difference that was statistically significant compared to docetaxel alone (p=0.03).

    Journal: International Journal of Oncology

    Article Title: Polysaccharide-K augments docetaxel-induced tumor suppression and antitumor immune response in an immunocompetent murine model of human prostate cancer

    doi: 10.3892/ijo.2011.1292

    Figure Lengend Snippet: WBC in tumor bearing mice treated with PSK and docetaxel. Groups of mice (n=7) bearing established TRAMP-C2 tumors (12 days) were treated with either oral saline, subtherapeutic dose of docetaxel (5 mg/kg twice weekly), oral gavage of PSK (300 mg/kg), or a combination of docetaxel and PSK. WBC were assessed at the end of the experiment (mean ± SEM). Docetaxel alone induced a 25% reduction in WBC. Addition of PSK decreased the docetaxel-suppressive effect by ~50%, a difference that was statistically significant compared to docetaxel alone (p=0.03).

    Article Snippet: PSK was donated by Kureha Chemical Industry (Tokyo, Japan) and docetaxel obtained from Sanofi-Aventis USA (Bridgewater, NJ).

    Techniques: Mouse Assay

    Antigen-specific responses and antigen cascade following vaccine plus docetaxel. CEA-Tg mice ( n = 3/group) were implanted s.c. with MC38-CEA + tumors on day 0. Mice were primed with rV-CEA/TRICOM on day 4 and boosted with rF-CEA/TRICOM on day 11. Both

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Antigen-specific responses and antigen cascade following vaccine plus docetaxel. CEA-Tg mice ( n = 3/group) were implanted s.c. with MC38-CEA + tumors on day 0. Mice were primed with rV-CEA/TRICOM on day 4 and boosted with rF-CEA/TRICOM on day 11. Both

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques: Mouse Assay

    Lack of effect of docetaxel on Treg function. C57BL/6 mice ( n = 5/group) were given 0.5 mg docetaxel i.p. on days -4, -2, and 0. Control mice received PBS only. Mice were sacrificed 1 day post-treatment. CD8 + cells from control mice were incubated with

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Lack of effect of docetaxel on Treg function. C57BL/6 mice ( n = 5/group) were given 0.5 mg docetaxel i.p. on days -4, -2, and 0. Control mice received PBS only. Mice were sacrificed 1 day post-treatment. CD8 + cells from control mice were incubated with

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques: Mouse Assay, Incubation

    Effect of docetaxel on splenic-cell populations and Treg function

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Effect of docetaxel on splenic-cell populations and Treg function

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques:

    Effect of docetaxel on splenic CD4 + and CD8 + T-cell cytokine production. C57BL/6 mice ( n = 5/group) were given 0.5 mg docetaxel i.p. on days -4, -2, and 0. Control mice received PBS only. Mice were sacrificed 1, 3, and 7 days post-treatment. A-E , splenocytes

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Effect of docetaxel on splenic CD4 + and CD8 + T-cell cytokine production. C57BL/6 mice ( n = 5/group) were given 0.5 mg docetaxel i.p. on days -4, -2, and 0. Control mice received PBS only. Mice were sacrificed 1, 3, and 7 days post-treatment. A-E , splenocytes

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques: Mouse Assay

    Antigen-specific T-cell responses combining vaccination plus docetaxel. A , CD4 + T-cell responses to foreign antigen . C57BL/6 mice ( n = 3/group) were primed on day 0 with rV-LacZ/TRICOM s.c. and boosted on day 14 with rF-LacZ/TRICOM s.c. Both vaccinations

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Antigen-specific T-cell responses combining vaccination plus docetaxel. A , CD4 + T-cell responses to foreign antigen . C57BL/6 mice ( n = 3/group) were primed on day 0 with rV-LacZ/TRICOM s.c. and boosted on day 14 with rF-LacZ/TRICOM s.c. Both vaccinations

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques: Mouse Assay

    Docetaxel given prior to recombinant viral vaccine can inhibit infection or transgene expression of susceptible cells. A , Murine tumor cells (MC38-CEA + ) were infected with rF-TRICOM (black bar). B , Cells were incubated with 1100 ng/mL (equivalent to clinical

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Docetaxel given prior to recombinant viral vaccine can inhibit infection or transgene expression of susceptible cells. A , Murine tumor cells (MC38-CEA + ) were infected with rF-TRICOM (black bar). B , Cells were incubated with 1100 ng/mL (equivalent to clinical

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques: Recombinant, Infection, Expressing, Incubation

    Increased antitumor activity of vaccine plus docetaxel. A , CEA-Tg mice ( n = 15/group) were implanted s.c. with MC38-CEA + tumors on day 0. Mice were primed with rV-CEA/TRICOM on day 4 and boosted with rF-CEA/TRICOM on day 11 (open arrows). Both vaccinations

    Journal:

    Article Title: Combination of Docetaxel and Recombinant Vaccine Enhances T-Cell Responses and Antitumor Activity

    doi: 10.1158/1078-0432.CCR-07-4025

    Figure Lengend Snippet: Increased antitumor activity of vaccine plus docetaxel. A , CEA-Tg mice ( n = 15/group) were implanted s.c. with MC38-CEA + tumors on day 0. Mice were primed with rV-CEA/TRICOM on day 4 and boosted with rF-CEA/TRICOM on day 11 (open arrows). Both vaccinations

    Article Snippet: Preparations of docetaxel (Taxotere™, kindly provided by Sanofi-Aventis, Bridgewater, NJ) were diluted to 10 mg/mL in 80% ethanol according to the manufacturer’s instructions, then further diluted to 5 mg/mL in sterile PBS.

    Techniques: Activity Assay, Mouse Assay

    NF 2 and RB 1 score as sensitizing candidates with vinorelbine. (A) Unique gene‐trap insertions in NF 2 and RB 1 in untreated ( n = 4, left panel), docetaxel‐treated ( n = 2, panel in the center), and vinorelbine‐treated ( n = 2, right panel) conditions. A representative example of each screening condition is shown including the P ‐value comparing the depicted drug‐treated replicate to the depicted untreated replicate, determined by Fisher's exact t ‐tests. Y ‐axis represents the sense to antisense integration ratio, while log 10 of sense/total number of insertions is plotted on the x ‐axis. Loss of RB 1 scored at times with a ratio > 0.5 in untreated and docetaxel‐treated screens, indicating that disruptive sense integrations caused a survival benefit. In vinorelbine screens, a ratio

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: NF 2 and RB 1 score as sensitizing candidates with vinorelbine. (A) Unique gene‐trap insertions in NF 2 and RB 1 in untreated ( n = 4, left panel), docetaxel‐treated ( n = 2, panel in the center), and vinorelbine‐treated ( n = 2, right panel) conditions. A representative example of each screening condition is shown including the P ‐value comparing the depicted drug‐treated replicate to the depicted untreated replicate, determined by Fisher's exact t ‐tests. Y ‐axis represents the sense to antisense integration ratio, while log 10 of sense/total number of insertions is plotted on the x ‐axis. Loss of RB 1 scored at times with a ratio > 0.5 in untreated and docetaxel‐treated screens, indicating that disruptive sense integrations caused a survival benefit. In vinorelbine screens, a ratio

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques:

    Identification of FBXW 7 mutation as a genetic vulnerability to vinorelbine. (A) Unique gene‐trap insertions in FBXW 7 in untreated ( n = 4, left panel), docetaxel‐treated ( n = 2, panel in the center), and vinorelbine‐treated ( n = 2, right panel) conditions. A representative example of each screening condition is shown including the P ‐value comparing the depicted drug‐treated replicate to the depicted untreated replicate, determined by Fisher's exact t ‐tests. Y ‐axis represents the sense to antisense integration ratio, while log 10 of sense/total number of insertions is plotted on the x ‐axis. Loss of FBXW 7 scored with a ratio > 0.5 in untreated conditions and docetaxel‐treated screens, indicating that disruptive sense integrations caused a survival benefit. For vinorelbine treatment, loss of FBXW 7 resulted in a disadvantage for survival, represented by a ratio

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: Identification of FBXW 7 mutation as a genetic vulnerability to vinorelbine. (A) Unique gene‐trap insertions in FBXW 7 in untreated ( n = 4, left panel), docetaxel‐treated ( n = 2, panel in the center), and vinorelbine‐treated ( n = 2, right panel) conditions. A representative example of each screening condition is shown including the P ‐value comparing the depicted drug‐treated replicate to the depicted untreated replicate, determined by Fisher's exact t ‐tests. Y ‐axis represents the sense to antisense integration ratio, while log 10 of sense/total number of insertions is plotted on the x ‐axis. Loss of FBXW 7 scored with a ratio > 0.5 in untreated conditions and docetaxel‐treated screens, indicating that disruptive sense integrations caused a survival benefit. For vinorelbine treatment, loss of FBXW 7 resulted in a disadvantage for survival, represented by a ratio

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques: Mutagenesis

    Inhibition of Mps1 by reversine protects Δ FBXW 7 cells from vinorelbine temporarily, and vinorelbine treatment causes a shift toward polyploidy in Δ FBXW 7 cells. (A) Equal numbers of cells were treated as indicated. Reversine treatment for 24 h could protect Δ FBXW 7 cells from vinorelbine hypersensitivity. Longer exposure to reversine resulted in reduced survival and increased toxicity of combined treatment with vinorelbine in all three genotypes. Bar plot shows mean quantification of three biological replicates with SEM ; * P = 0.01–0.05. (B) DNA profiles measured by DAPI intensity after 18 h of treatment with 1.3× IC 50 concentrations of docetaxel or vinorelbine. Note that Δ FBXW 7 cells shift toward G2/M and polyploidy upon vinorelbine exposure ( n = 2).

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: Inhibition of Mps1 by reversine protects Δ FBXW 7 cells from vinorelbine temporarily, and vinorelbine treatment causes a shift toward polyploidy in Δ FBXW 7 cells. (A) Equal numbers of cells were treated as indicated. Reversine treatment for 24 h could protect Δ FBXW 7 cells from vinorelbine hypersensitivity. Longer exposure to reversine resulted in reduced survival and increased toxicity of combined treatment with vinorelbine in all three genotypes. Bar plot shows mean quantification of three biological replicates with SEM ; * P = 0.01–0.05. (B) DNA profiles measured by DAPI intensity after 18 h of treatment with 1.3× IC 50 concentrations of docetaxel or vinorelbine. Note that Δ FBXW 7 cells shift toward G2/M and polyploidy upon vinorelbine exposure ( n = 2).

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques: Inhibition

    Less Δ FBXW 7 cells are in mitosis after 18 h of vinorelbine treatment. Representative images of antitubulin immunofluorescence staining after 18 h of treatment with 0.8× IC 50 of vinorelbine or docetaxel. Cells in prometaphase to anaphase stage of mitosis, indicated with arrows, were counted in 4× 16 adjacent 100× power fields as percentage of total number of cells. Scale bar represents 10 μm. Mitotic cells in the three indicated cell lines, normalized to percentage of mitotic cells without treatment, of one experiment are quantified in the bar blot with SD . Note the significant lower number of mitotic cells in the Δ FBXW 7 cell line. * P = 0.01–0.05, n.s. = not significant, n = 2.

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: Less Δ FBXW 7 cells are in mitosis after 18 h of vinorelbine treatment. Representative images of antitubulin immunofluorescence staining after 18 h of treatment with 0.8× IC 50 of vinorelbine or docetaxel. Cells in prometaphase to anaphase stage of mitosis, indicated with arrows, were counted in 4× 16 adjacent 100× power fields as percentage of total number of cells. Scale bar represents 10 μm. Mitotic cells in the three indicated cell lines, normalized to percentage of mitotic cells without treatment, of one experiment are quantified in the bar blot with SD . Note the significant lower number of mitotic cells in the Δ FBXW 7 cell line. * P = 0.01–0.05, n.s. = not significant, n = 2.

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques: Immunofluorescence, Staining

    Δ FBXW 7 cells die more frequently in mitosis, undergo multinucleation, and show an increased prometaphase to metaphase time interval. (A) Representative images of H2B‐ GFP ‐tagged diploid wild‐type and Δ FBXW 7 cells during time‐lapse microscopy with or without MTA treatment are shown on a time line starting at the last image before entry into mitosis. Under untreated conditions, no phenotypic mitotic alterations were observed in both wild‐type and Δ FBXW 7 cells. Vinorelbine treatment caused repeatedly formation of metaphase plates in both cell lines, in the course of which Δ FBXW 7 cells died more frequently compared to wild‐type cells. Docetaxel treatment caused an increase in multipolar spindle configurations, as shown in the examples in wild‐type and Δ FBXW 7 cells. Scale bar represents 10 μm. Scatter plots show quantification of four time‐lapse experiments. Treatment with either vinorelbine or docetaxel caused a significant increase in duration of mitosis (from prometaphase to anaphase) in both cell lines. No significant difference between wild‐type and Δ FBXW 7 cells was seen in overall duration of mitosis. However, a significant prolonged prometaphase to metaphase time interval was observed in Δ FBXW 7 compared to wild‐type cells upon vinorelbine treatment, and a shorter duration of metaphase to anaphase transition. **** P

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: Δ FBXW 7 cells die more frequently in mitosis, undergo multinucleation, and show an increased prometaphase to metaphase time interval. (A) Representative images of H2B‐ GFP ‐tagged diploid wild‐type and Δ FBXW 7 cells during time‐lapse microscopy with or without MTA treatment are shown on a time line starting at the last image before entry into mitosis. Under untreated conditions, no phenotypic mitotic alterations were observed in both wild‐type and Δ FBXW 7 cells. Vinorelbine treatment caused repeatedly formation of metaphase plates in both cell lines, in the course of which Δ FBXW 7 cells died more frequently compared to wild‐type cells. Docetaxel treatment caused an increase in multipolar spindle configurations, as shown in the examples in wild‐type and Δ FBXW 7 cells. Scale bar represents 10 μm. Scatter plots show quantification of four time‐lapse experiments. Treatment with either vinorelbine or docetaxel caused a significant increase in duration of mitosis (from prometaphase to anaphase) in both cell lines. No significant difference between wild‐type and Δ FBXW 7 cells was seen in overall duration of mitosis. However, a significant prolonged prometaphase to metaphase time interval was observed in Δ FBXW 7 compared to wild‐type cells upon vinorelbine treatment, and a shorter duration of metaphase to anaphase transition. **** P

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques: Time-lapse Microscopy

    Identification of ABCB 1 validates the concept of the screens, while docetaxel and vinorelbine display different genetic vulnerabilities. (A) Unique gene‐trap insertions in ABCB 1 in untreated ( n = 4, left panel), docetaxel‐treated ( n = 2, panel in the center), and vinorelbine‐treated ( n = 2, right panel) conditions. A representative example of each screening condition is shown including the P ‐value comparing the depicted drug‐treated replicate to the depicted untreated replicate, determined by Fisher's exact t ‐tests. Y ‐axis represents the sense to antisense integration ratio, while log 10 of sense/total number of insertions is plotted on the x ‐axis. Loss of ABCB 1 was neutral in regard to cell survival without drug treatment, represented by a 0.5 sense to antisense ratio. Upon docetaxel or vinorelbine treatment, ABCB 1 was depleted for sense insertions (ratio

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: Identification of ABCB 1 validates the concept of the screens, while docetaxel and vinorelbine display different genetic vulnerabilities. (A) Unique gene‐trap insertions in ABCB 1 in untreated ( n = 4, left panel), docetaxel‐treated ( n = 2, panel in the center), and vinorelbine‐treated ( n = 2, right panel) conditions. A representative example of each screening condition is shown including the P ‐value comparing the depicted drug‐treated replicate to the depicted untreated replicate, determined by Fisher's exact t ‐tests. Y ‐axis represents the sense to antisense integration ratio, while log 10 of sense/total number of insertions is plotted on the x ‐axis. Loss of ABCB 1 was neutral in regard to cell survival without drug treatment, represented by a 0.5 sense to antisense ratio. Upon docetaxel or vinorelbine treatment, ABCB 1 was depleted for sense insertions (ratio

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques:

    Layout of the insertional mutagenesis haploid screens. (A) Wild‐type HAP 1 cells were gene‐trap mutagenized, exposed to 16 n m vinorelbine or 7 n m docetaxel, and subsequently allowed to recover until day 10. Genomic DNA of 3 × 10 7 cells with 1n DNA content was extracted; insertion sites were amplified by LAM ‐ PCR before sequencing, mapping to the human genome, and normalizing to untreated cultured control datasets. Genes without effect on cellular fitness will have approximately equal numbers of disruptive sense and nondisruptive antisense integrations (ratio 0.5, neutral gene). Genes in which mutations increase survival of the cell will score with a higher proportion of sense integrations. Fewer sense integrations compared to antisense integrations will be counted in hypersensitivity genes. (B) Illustrated in a simplified fashion, intronic gene‐trap sense integration in relation to the transcriptional direction of a gene is disruptive, whereas antisense integration does not affect the function of a transcript.

    Journal: Molecular Oncology

    Article Title: Haploid genetic screens identify genetic vulnerabilities to microtubule‐targeting agents

    doi: 10.1002/1878-0261.12307

    Figure Lengend Snippet: Layout of the insertional mutagenesis haploid screens. (A) Wild‐type HAP 1 cells were gene‐trap mutagenized, exposed to 16 n m vinorelbine or 7 n m docetaxel, and subsequently allowed to recover until day 10. Genomic DNA of 3 × 10 7 cells with 1n DNA content was extracted; insertion sites were amplified by LAM ‐ PCR before sequencing, mapping to the human genome, and normalizing to untreated cultured control datasets. Genes without effect on cellular fitness will have approximately equal numbers of disruptive sense and nondisruptive antisense integrations (ratio 0.5, neutral gene). Genes in which mutations increase survival of the cell will score with a higher proportion of sense integrations. Fewer sense integrations compared to antisense integrations will be counted in hypersensitivity genes. (B) Illustrated in a simplified fashion, intronic gene‐trap sense integration in relation to the transcriptional direction of a gene is disruptive, whereas antisense integration does not affect the function of a transcript.

    Article Snippet: 108 mutagenized HAP1 cells were seeded in 14 T175 cell culture flasks (Corning, New York, NY, USA), treated 24 h after seeding with either 4.5× IC50 of docetaxel (7 nm , Taxotere; Sanofi Aventis, Paris, France) or 6.5× IC50 of vinorelbine (16 nm , Vinorelbine; Actavis, Luxembourg, Luxembourg), as determined in nonmutagenized wild‐type HAP1 cells (see section ).

    Techniques: Mutagenesis, Amplification, Laser Capture Microdissection, Polymerase Chain Reaction, Sequencing, Cell Culture

    MiR-27b regulates the tumour seeding ability of breast cancer cells. ( a ) Flow cytometric analyses of MCF7-luc Zs-DR-27bs cells transfected with LNA-NC or LNA-miR-27b. ( b ) Overview of the method used to analyse the CSC properties of docetaxel-treated MCF7-luc cell derivatives. ( c ) Bioluminescent images of tumours in NOD/SCID mice injected with docetaxel (DOC)-treated MCF7-luc cell derivatives. Representative images are shown for each cohort. ( d ) The numbers of animals with detectable tumours in the groups injected with the MCF7-luc cell derivatives.

    Journal: Nature Communications

    Article Title: Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1

    doi: 10.1038/ncomms8318

    Figure Lengend Snippet: MiR-27b regulates the tumour seeding ability of breast cancer cells. ( a ) Flow cytometric analyses of MCF7-luc Zs-DR-27bs cells transfected with LNA-NC or LNA-miR-27b. ( b ) Overview of the method used to analyse the CSC properties of docetaxel-treated MCF7-luc cell derivatives. ( c ) Bioluminescent images of tumours in NOD/SCID mice injected with docetaxel (DOC)-treated MCF7-luc cell derivatives. Representative images are shown for each cohort. ( d ) The numbers of animals with detectable tumours in the groups injected with the MCF7-luc cell derivatives.

    Article Snippet: Metformin (1,1-dimethylbiguanide hydrochloride; Sigma-Aldrich) was used at 0.1–100 mM and docetaxel (Sanofi Aventis) was used at 1–10 nM.

    Techniques: Flow Cytometry, Transfection, Mouse Assay, Injection

    MiR-27b regulates the resistance of breast cancer cells to docetaxel. ( a ) Overview of the method used to establish miR-27b knockdown MCF7-luc (MCF7-luc anti-miR-27b) cells. ( b , c ) Dose–response curves of MCF7-luc anti-NC, MCF7-luc anti-miR-27b and MCF7-luc miR-27b o.e. cells treated with docetaxel. Cell viability was normalized to that of the corresponding cells treated with dimethylsulphoxide (DMSO). The red dashed line indicates the IC 50 value. Data are represented as the mean±s.d. of n =3 replicates. ( d ) Morphologies of the MCF7-luc anti-NC, MCF7-luc miR-27b o.e. and MCF7-luc anti-miR-27b cells. Scale bar, 100 μm. ( e ) Flow cytometric analyses of the SP fraction of MCF7-luc derivatives in the presence and absence of Ko143. ( f ) Quantification of the SP fraction of MCF7-luc derivatives. The SP fraction was determined as the difference between the level of Hoechst 33342 staining in the presence and absence of Ko143. Data are represented as the mean±s.d. of n =3 replicates. Statistical significance was determined by Student's t -test.

    Journal: Nature Communications

    Article Title: Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1

    doi: 10.1038/ncomms8318

    Figure Lengend Snippet: MiR-27b regulates the resistance of breast cancer cells to docetaxel. ( a ) Overview of the method used to establish miR-27b knockdown MCF7-luc (MCF7-luc anti-miR-27b) cells. ( b , c ) Dose–response curves of MCF7-luc anti-NC, MCF7-luc anti-miR-27b and MCF7-luc miR-27b o.e. cells treated with docetaxel. Cell viability was normalized to that of the corresponding cells treated with dimethylsulphoxide (DMSO). The red dashed line indicates the IC 50 value. Data are represented as the mean±s.d. of n =3 replicates. ( d ) Morphologies of the MCF7-luc anti-NC, MCF7-luc miR-27b o.e. and MCF7-luc anti-miR-27b cells. Scale bar, 100 μm. ( e ) Flow cytometric analyses of the SP fraction of MCF7-luc derivatives in the presence and absence of Ko143. ( f ) Quantification of the SP fraction of MCF7-luc derivatives. The SP fraction was determined as the difference between the level of Hoechst 33342 staining in the presence and absence of Ko143. Data are represented as the mean±s.d. of n =3 replicates. Statistical significance was determined by Student's t -test.

    Article Snippet: Metformin (1,1-dimethylbiguanide hydrochloride; Sigma-Aldrich) was used at 0.1–100 mM and docetaxel (Sanofi Aventis) was used at 1–10 nM.

    Techniques: Flow Cytometry, Staining

    Downregulation of miR-27b is associated with the generation of CD44 high /CD24 low fraction. ( a ) Flow cytometry analyses of the CD44 high /CD24 low population and Zs-DR expression in MCF7-luc Zs-DR-27bs and its tumorigenic cells (MCF7-luc-ZT1 cells) in adherent and mammosphere culture conditions. ( b ) Flow cytometry analyses of the CD44 high /CD24 low and GFP high populations of ZR75-1-luc anti-miR-27b cells treated with docetaxel (DOC).

    Journal: Nature Communications

    Article Title: Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1

    doi: 10.1038/ncomms8318

    Figure Lengend Snippet: Downregulation of miR-27b is associated with the generation of CD44 high /CD24 low fraction. ( a ) Flow cytometry analyses of the CD44 high /CD24 low population and Zs-DR expression in MCF7-luc Zs-DR-27bs and its tumorigenic cells (MCF7-luc-ZT1 cells) in adherent and mammosphere culture conditions. ( b ) Flow cytometry analyses of the CD44 high /CD24 low and GFP high populations of ZR75-1-luc anti-miR-27b cells treated with docetaxel (DOC).

    Article Snippet: Metformin (1,1-dimethylbiguanide hydrochloride; Sigma-Aldrich) was used at 0.1–100 mM and docetaxel (Sanofi Aventis) was used at 1–10 nM.

    Techniques: Flow Cytometry, Cytometry, Expressing

    Functional analysis of ENPP1 in MCF7-luc cells. ( a ) Flow cytometric analysis of the SP fractions of MCF7-luc cells overexpressing ENPP1-MF or GFP as a control, in the presence and absence of Ko143. ( b ) Quantification of the SP fractions shown in a , determined as the difference between the level of Hoechst 33342 staining in the presence and absence of Ko143. Data are represented as the mean±s.d. of n =3 replicates. ( c ) Flow cytometric analysis showing the cell surface localization of ABCG2 in the indicated 293T co-transfectants. ( d ) Flow cytometric analyses of the cell surface localization of ABCG2 in MCF7-luc anti-miR-27b cells transfected with a control (shNC) or ENPP1-specific (shENPP1) shRNA. ( e ) Dose–response curves of docetaxel-treated MCF7-luc anti-miR-27b-DR cells transfected with shNC or shENPP1. Cell viability was normalized to that of the corresponding cells treated with dimethylsulphoxide (DMSO). The red dashed line indicates the IC 50 value. Data are represented as the mean±s.d. of n =3 replicates. ( f ) Proximity ligation assay using MCF7-luc anti-NC or MCF7-luc anti-miR-27b cells transiently expressing ABCG2-HA. Scale bar, 50 μm. ( g ) In vitro binding assay using C-terminally Flag-tagged GFP or C-terminally Myc- and Flag-tagged ENPP1 purified from 293T cells and C-terminally HA-tagged ABCG2 purified from Sf21 insect cell extracts.

    Journal: Nature Communications

    Article Title: Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1

    doi: 10.1038/ncomms8318

    Figure Lengend Snippet: Functional analysis of ENPP1 in MCF7-luc cells. ( a ) Flow cytometric analysis of the SP fractions of MCF7-luc cells overexpressing ENPP1-MF or GFP as a control, in the presence and absence of Ko143. ( b ) Quantification of the SP fractions shown in a , determined as the difference between the level of Hoechst 33342 staining in the presence and absence of Ko143. Data are represented as the mean±s.d. of n =3 replicates. ( c ) Flow cytometric analysis showing the cell surface localization of ABCG2 in the indicated 293T co-transfectants. ( d ) Flow cytometric analyses of the cell surface localization of ABCG2 in MCF7-luc anti-miR-27b cells transfected with a control (shNC) or ENPP1-specific (shENPP1) shRNA. ( e ) Dose–response curves of docetaxel-treated MCF7-luc anti-miR-27b-DR cells transfected with shNC or shENPP1. Cell viability was normalized to that of the corresponding cells treated with dimethylsulphoxide (DMSO). The red dashed line indicates the IC 50 value. Data are represented as the mean±s.d. of n =3 replicates. ( f ) Proximity ligation assay using MCF7-luc anti-NC or MCF7-luc anti-miR-27b cells transiently expressing ABCG2-HA. Scale bar, 50 μm. ( g ) In vitro binding assay using C-terminally Flag-tagged GFP or C-terminally Myc- and Flag-tagged ENPP1 purified from 293T cells and C-terminally HA-tagged ABCG2 purified from Sf21 insect cell extracts.

    Article Snippet: Metformin (1,1-dimethylbiguanide hydrochloride; Sigma-Aldrich) was used at 0.1–100 mM and docetaxel (Sanofi Aventis) was used at 1–10 nM.

    Techniques: Functional Assay, Flow Cytometry, Staining, Transfection, shRNA, Proximity Ligation Assay, Expressing, Hemagglutination Assay, In Vitro, Binding Assay, Purification

    MiR-27b is involved in the drug resistance and tumorigenicity of breast cancer cells. ( a ) Relative miR-27b expression levels in normal and luminal-type breast cancer tissues. Expression levels were normalized to those of RNU6B . Differences between groups were analysed using unpaired t -tests. ( b ) Schematic illustration of the miR-27b sensor construct used in the experiments shown in c – f . ( c ) Expression of Zs-DR 48 h after transfection of MCF7-luc Zs-DR-27bs cells with a negative control or miR-27b-specific LNA. Scale bar, 100 μm. ( d ) Flow cytometric analyses of Zs-DR expression 48 h after treatment of MCF7-luc Zs-DR-27bs cells with dimethylsulphoxide (DMSO) or docetaxel (DOC). ( e ) Isolation of tumourigenic MCF7-luc Zs-DR-27bs cells. After transplantation of MCF7-luc Zs-DR-27bs cells into 5-week-old NOD/SCID mice, the cells were isolated and cultivated in vitro . ( f ) Flow cytometric analyses of Zs-DR expression in tumourigenic MCF7-luc Zs-DR-27bs cells isolated from the mice described in e .

    Journal: Nature Communications

    Article Title: Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1

    doi: 10.1038/ncomms8318

    Figure Lengend Snippet: MiR-27b is involved in the drug resistance and tumorigenicity of breast cancer cells. ( a ) Relative miR-27b expression levels in normal and luminal-type breast cancer tissues. Expression levels were normalized to those of RNU6B . Differences between groups were analysed using unpaired t -tests. ( b ) Schematic illustration of the miR-27b sensor construct used in the experiments shown in c – f . ( c ) Expression of Zs-DR 48 h after transfection of MCF7-luc Zs-DR-27bs cells with a negative control or miR-27b-specific LNA. Scale bar, 100 μm. ( d ) Flow cytometric analyses of Zs-DR expression 48 h after treatment of MCF7-luc Zs-DR-27bs cells with dimethylsulphoxide (DMSO) or docetaxel (DOC). ( e ) Isolation of tumourigenic MCF7-luc Zs-DR-27bs cells. After transplantation of MCF7-luc Zs-DR-27bs cells into 5-week-old NOD/SCID mice, the cells were isolated and cultivated in vitro . ( f ) Flow cytometric analyses of Zs-DR expression in tumourigenic MCF7-luc Zs-DR-27bs cells isolated from the mice described in e .

    Article Snippet: Metformin (1,1-dimethylbiguanide hydrochloride; Sigma-Aldrich) was used at 0.1–100 mM and docetaxel (Sanofi Aventis) was used at 1–10 nM.

    Techniques: Expressing, Construct, Transfection, Negative Control, Flow Cytometry, Isolation, Transplantation Assay, Mouse Assay, In Vitro

    ENPP1 is a substrate of the 26S proteasome. ( a ) ENPP1 expression in MCF7-luc cells stably expressing ENPP1-MF or anti-NC as a control, detected by qRT–PCR. Expression levels were normalized to those of GAPDH and data are represented as the mean±s.d. of n =3 replicates. The P -values were calculated by Student's t -test. ( b ) A schematic illustration of the approach used in the experiments shown in c – e . ( c ) Immunoblot analyses of ENPP1 expression in MCF7-luc anti-NC and MCF7-luc ENPP1-MF cells treated with or without MG132 for 24 h. ( d ) Immunoblot analyses of ENPP1 in MCF7-luc ENPP1-MF cells grown under adherent (Ad) or mammosphere (Ma) culture conditions for the indicated times. ( e ) ENPP1 expression in the indicated MCF7-luc derivatives grown under adherent (Ad) or mammosphere (Ma) culture conditions. ( f ) Growth of the indicated MCF7-luc cell derivatives. An MTT assay was performed to determine the numbers of cells at each time point. Data are represented as the mean±s.d. of n =3 replicates. The P -values were calculated by Student's t -test. ( g ) Immunofluorescent detection of ENPP1-MF and GFP in MCF7-luc ENPP1-MF cells in paraffin-embedded sections of primary tumour xenografts. The nuclei were stained with 4,6-diamidino-2-phenylindole (DAPI). Scale bar, 50 μm. ( h ) Bioluminescent images of tumours in NOD/SCID mice injected with MCF7-luc shENPP1 cells that were treated with or without docetaxel (DOC). Alternatively, the mice were injected with MCF7-luc Zs-DR-27bs cells as a technical control. Representative images are shown for each cohort.

    Journal: Nature Communications

    Article Title: Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1

    doi: 10.1038/ncomms8318

    Figure Lengend Snippet: ENPP1 is a substrate of the 26S proteasome. ( a ) ENPP1 expression in MCF7-luc cells stably expressing ENPP1-MF or anti-NC as a control, detected by qRT–PCR. Expression levels were normalized to those of GAPDH and data are represented as the mean±s.d. of n =3 replicates. The P -values were calculated by Student's t -test. ( b ) A schematic illustration of the approach used in the experiments shown in c – e . ( c ) Immunoblot analyses of ENPP1 expression in MCF7-luc anti-NC and MCF7-luc ENPP1-MF cells treated with or without MG132 for 24 h. ( d ) Immunoblot analyses of ENPP1 in MCF7-luc ENPP1-MF cells grown under adherent (Ad) or mammosphere (Ma) culture conditions for the indicated times. ( e ) ENPP1 expression in the indicated MCF7-luc derivatives grown under adherent (Ad) or mammosphere (Ma) culture conditions. ( f ) Growth of the indicated MCF7-luc cell derivatives. An MTT assay was performed to determine the numbers of cells at each time point. Data are represented as the mean±s.d. of n =3 replicates. The P -values were calculated by Student's t -test. ( g ) Immunofluorescent detection of ENPP1-MF and GFP in MCF7-luc ENPP1-MF cells in paraffin-embedded sections of primary tumour xenografts. The nuclei were stained with 4,6-diamidino-2-phenylindole (DAPI). Scale bar, 50 μm. ( h ) Bioluminescent images of tumours in NOD/SCID mice injected with MCF7-luc shENPP1 cells that were treated with or without docetaxel (DOC). Alternatively, the mice were injected with MCF7-luc Zs-DR-27bs cells as a technical control. Representative images are shown for each cohort.

    Article Snippet: Metformin (1,1-dimethylbiguanide hydrochloride; Sigma-Aldrich) was used at 0.1–100 mM and docetaxel (Sanofi Aventis) was used at 1–10 nM.

    Techniques: Expressing, Stable Transfection, Quantitative RT-PCR, MTT Assay, Staining, Mouse Assay, Injection

    Effect of exogenous expression of MRP2 on the cytotoxicity ( a ) and intracellular accumulation ( b ) of docetaxel. ( a ) Vector-transfected MDCKII cells (closed circles) or MRP2-expressing MDCKII cells (open circles) were incubated for 96 h with various concentrations (0.01–100 nmol/l) of docetaxel. Cell survival was quantified by the MTT assay as described in the Methods. ( b ) Cell lines were incubated for 24 h with 1 nmol/l of docetaxel, and the volume uptake of docetaxel into these cells was determined. Each point and bar represents the mean ± SE ( n = 3). * P

    Journal: CPT: Pharmacometrics & Systems Pharmacology

    Article Title: Kinetic Interpretation of the Importance of OATP1B3 and MRP2 in Docetaxel-Induced Hematopoietic Toxicity

    doi: 10.1038/psp.2014.23

    Figure Lengend Snippet: Effect of exogenous expression of MRP2 on the cytotoxicity ( a ) and intracellular accumulation ( b ) of docetaxel. ( a ) Vector-transfected MDCKII cells (closed circles) or MRP2-expressing MDCKII cells (open circles) were incubated for 96 h with various concentrations (0.01–100 nmol/l) of docetaxel. Cell survival was quantified by the MTT assay as described in the Methods. ( b ) Cell lines were incubated for 24 h with 1 nmol/l of docetaxel, and the volume uptake of docetaxel into these cells was determined. Each point and bar represents the mean ± SE ( n = 3). * P

    Article Snippet: Unlabeled E2 17βG, E1 S, and CCK-8 were purchased from Sigma-Aldrich (St Louis, MO), and docetaxel was purchased from LC Laboratories (Woburn, MA).

    Techniques: Expressing, Plasmid Preparation, Transfection, Incubation, MTT Assay

    Schematic diagram of the PK/PD model describing docetaxel-induced neutropenia. ( a ) A three-compartment model was used to describe the pharmacokinetics of docetaxel. ( b ) The model of Friberg et al . was used to estimate the neutrophil count. 17 ( c ) Neutrophil concentration profile after docetaxel administration in five individuals selected from subjects with SLCO1B3 and ABCC2 diplotypes of wt/wt and wt/wt. Broken lines show borders between non-ADR, neutropenia grade 1/2, and neutropenia grade 3/4. Non-ADR, nonadverse drug reaction; PD, pharmacodynamic; PK, pharmacokinetic.

    Journal: CPT: Pharmacometrics & Systems Pharmacology

    Article Title: Kinetic Interpretation of the Importance of OATP1B3 and MRP2 in Docetaxel-Induced Hematopoietic Toxicity

    doi: 10.1038/psp.2014.23

    Figure Lengend Snippet: Schematic diagram of the PK/PD model describing docetaxel-induced neutropenia. ( a ) A three-compartment model was used to describe the pharmacokinetics of docetaxel. ( b ) The model of Friberg et al . was used to estimate the neutrophil count. 17 ( c ) Neutrophil concentration profile after docetaxel administration in five individuals selected from subjects with SLCO1B3 and ABCC2 diplotypes of wt/wt and wt/wt. Broken lines show borders between non-ADR, neutropenia grade 1/2, and neutropenia grade 3/4. Non-ADR, nonadverse drug reaction; PD, pharmacodynamic; PK, pharmacokinetic.

    Article Snippet: Unlabeled E2 17βG, E1 S, and CCK-8 were purchased from Sigma-Aldrich (St Louis, MO), and docetaxel was purchased from LC Laboratories (Woburn, MA).

    Techniques: Concentration Assay

    Inhibitory effect of docetaxel on granulocyte colony–stimulating factor (G-CSF)-induced colony formation. ( a ) Bone marrow cells obtained from Sprague Dawley rats (open circles) and Eisai hyperbilirubinemic rats (closed circles) were incubated for 6 days with various concentrations (0–5 ng/ml) of docetaxel. ( b ) Bone marrow cells derived from Sprague Dawley rats were also incubated for 6 days with various concentrations (0–5 ng/ml) of docetaxel in the presence (closed circles) or absence (open circles) of 50 µmol/l MK571. The number of colonies induced by G-CSF was counted. Each point represents the mean ± SE ( n = 9).* P

    Journal: CPT: Pharmacometrics & Systems Pharmacology

    Article Title: Kinetic Interpretation of the Importance of OATP1B3 and MRP2 in Docetaxel-Induced Hematopoietic Toxicity

    doi: 10.1038/psp.2014.23

    Figure Lengend Snippet: Inhibitory effect of docetaxel on granulocyte colony–stimulating factor (G-CSF)-induced colony formation. ( a ) Bone marrow cells obtained from Sprague Dawley rats (open circles) and Eisai hyperbilirubinemic rats (closed circles) were incubated for 6 days with various concentrations (0–5 ng/ml) of docetaxel. ( b ) Bone marrow cells derived from Sprague Dawley rats were also incubated for 6 days with various concentrations (0–5 ng/ml) of docetaxel in the presence (closed circles) or absence (open circles) of 50 µmol/l MK571. The number of colonies induced by G-CSF was counted. Each point represents the mean ± SE ( n = 9).* P

    Article Snippet: Unlabeled E2 17βG, E1 S, and CCK-8 were purchased from Sigma-Aldrich (St Louis, MO), and docetaxel was purchased from LC Laboratories (Woburn, MA).

    Techniques: Incubation, Derivative Assay

    In vitro uptake of docetaxel into HEK293 cells expressing solute carrier transporters expressed in human hepatocytes and into human hepatocytes. ( a ) The uptake of docetaxel (1 µmol/l) into OATP-, OAT-, OCT1-, and NTCP–expressing HEK293 cells and vector-transfected control cells (vector) was measured at 5 min. ( b ) Saturation of the docetaxel uptake in OATP1B3-expressing HEK293 cells was also investigated. The solid line is a fitted curve calculated by nonlinear regression analysis based on Eq. 1, as described in the Methods. ( c ) Concentration (0.1 µmol/l and 2 mmol/l)-dependent uptake of estrone-3-sulfate (E 1 S) in OATP1B1-expressing HEK293 cells, ( d ) inhibitory effects of 2 mmol/l estradiol-17β-glucuronide (E 2 17βG) and E 1 S on the uptake of cholecystokinin octapeptide (CCK-8) in OATP1B3-expressing HEK293 cells, and ( e ) docetaxel uptake into cryopreserved human hepatocytes in the presence of E 1 S and E 2 17βG were investigated. These uptake assays were carried out in the presence of 3% human serum albumin. Each bar represents the mean ± SE ( n = 3). * P

    Journal: CPT: Pharmacometrics & Systems Pharmacology

    Article Title: Kinetic Interpretation of the Importance of OATP1B3 and MRP2 in Docetaxel-Induced Hematopoietic Toxicity

    doi: 10.1038/psp.2014.23

    Figure Lengend Snippet: In vitro uptake of docetaxel into HEK293 cells expressing solute carrier transporters expressed in human hepatocytes and into human hepatocytes. ( a ) The uptake of docetaxel (1 µmol/l) into OATP-, OAT-, OCT1-, and NTCP–expressing HEK293 cells and vector-transfected control cells (vector) was measured at 5 min. ( b ) Saturation of the docetaxel uptake in OATP1B3-expressing HEK293 cells was also investigated. The solid line is a fitted curve calculated by nonlinear regression analysis based on Eq. 1, as described in the Methods. ( c ) Concentration (0.1 µmol/l and 2 mmol/l)-dependent uptake of estrone-3-sulfate (E 1 S) in OATP1B1-expressing HEK293 cells, ( d ) inhibitory effects of 2 mmol/l estradiol-17β-glucuronide (E 2 17βG) and E 1 S on the uptake of cholecystokinin octapeptide (CCK-8) in OATP1B3-expressing HEK293 cells, and ( e ) docetaxel uptake into cryopreserved human hepatocytes in the presence of E 1 S and E 2 17βG were investigated. These uptake assays were carried out in the presence of 3% human serum albumin. Each bar represents the mean ± SE ( n = 3). * P

    Article Snippet: Unlabeled E2 17βG, E1 S, and CCK-8 were purchased from Sigma-Aldrich (St Louis, MO), and docetaxel was purchased from LC Laboratories (Woburn, MA).

    Techniques: In Vitro, Expressing, Plasmid Preparation, Transfection, Concentration Assay, CCK-8 Assay

    Release of docetaxel from DTX-loaded PLGA and PLGA–PEG NP formulations (1, 6, and 12 h, followed by 1, 2, 3, 4, and 5 days). Abbreviations: PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Journal: International Journal of Nanomedicine

    Article Title: Docetaxel-loaded PLGA and PLGA-PEG nanoparticles for intravenous application: pharmacokinetics and biodistribution profile

    doi: 10.2147/IJN.S121881

    Figure Lengend Snippet: Release of docetaxel from DTX-loaded PLGA and PLGA–PEG NP formulations (1, 6, and 12 h, followed by 1, 2, 3, 4, and 5 days). Abbreviations: PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Article Snippet: Docetaxel was purchased from LC Laboratories (Woburn, MA, USA).

    Techniques:

    Docetaxel concentration in mouse kidney versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Journal: International Journal of Nanomedicine

    Article Title: Docetaxel-loaded PLGA and PLGA-PEG nanoparticles for intravenous application: pharmacokinetics and biodistribution profile

    doi: 10.2147/IJN.S121881

    Figure Lengend Snippet: Docetaxel concentration in mouse kidney versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Article Snippet: Docetaxel was purchased from LC Laboratories (Woburn, MA, USA).

    Techniques: Concentration Assay, IV Injection

    Docetaxel concentration in mouse lung versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Journal: International Journal of Nanomedicine

    Article Title: Docetaxel-loaded PLGA and PLGA-PEG nanoparticles for intravenous application: pharmacokinetics and biodistribution profile

    doi: 10.2147/IJN.S121881

    Figure Lengend Snippet: Docetaxel concentration in mouse lung versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Article Snippet: Docetaxel was purchased from LC Laboratories (Woburn, MA, USA).

    Techniques: Concentration Assay, IV Injection

    Docetaxel concentration in mouse heart versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Journal: International Journal of Nanomedicine

    Article Title: Docetaxel-loaded PLGA and PLGA-PEG nanoparticles for intravenous application: pharmacokinetics and biodistribution profile

    doi: 10.2147/IJN.S121881

    Figure Lengend Snippet: Docetaxel concentration in mouse heart versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Article Snippet: Docetaxel was purchased from LC Laboratories (Woburn, MA, USA).

    Techniques: Concentration Assay, IV Injection

    DTX serum concentration versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Journal: International Journal of Nanomedicine

    Article Title: Docetaxel-loaded PLGA and PLGA-PEG nanoparticles for intravenous application: pharmacokinetics and biodistribution profile

    doi: 10.2147/IJN.S121881

    Figure Lengend Snippet: DTX serum concentration versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Article Snippet: Docetaxel was purchased from LC Laboratories (Woburn, MA, USA).

    Techniques: Concentration Assay, IV Injection

    Docetaxel concentration in mouse liver versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Journal: International Journal of Nanomedicine

    Article Title: Docetaxel-loaded PLGA and PLGA-PEG nanoparticles for intravenous application: pharmacokinetics and biodistribution profile

    doi: 10.2147/IJN.S121881

    Figure Lengend Snippet: Docetaxel concentration in mouse liver versus time after IV injection of different drug formulations at a dose of 5 mg/kg (n=4). Note: * Statistically significant difference between treatment groups. Abbreviations: IV, intravenous; PEG, poly(ethylene glycol); PLGA, poly(lactide- co -glycolide); DTX, docetaxel; NP, nanoparticle.

    Article Snippet: Docetaxel was purchased from LC Laboratories (Woburn, MA, USA).

    Techniques: Concentration Assay, IV Injection

    Inhibition of ABL kinases in the presence of docetaxel decreases cell proliferation and increases cell death in Kras G12D/+ ; p53 −/− driven lung tumors Treatments began 10 weeks after Adeno-Cre infection of Rosa26-fGFP; LSL-Kras G12D/+ ; p53 fl/fl mice. Mouse lungs were harvested at 12 weeks after Adeno-Cre infection. A. IHC for Ki67+ cells in sections of mouse lungs from mice treated with vehicle control, docetaxel, GNF5, or combination (docetaxel+GNF5) treatment showing a decrease in Ki67 staining, particularly in the combination therapy group. Scale bar = 50 μm. B. Quantification of IHC staining for Ki67 ( n = 5-7 tumors per group). C. Immunofluorescence staining for Ki67 showing a decrease in the percentage of Ki67+ (red) tumor cells (labeled with farnesylated GFP, green) in mice treated with combination therapy of GNF5 and docetaxel compared to vehicle treated mice. Scale bar = 150 μm. D. Immunoblotting of lysates showed a decrease in Ki67 expression and increase in cleaved caspase 3 expression in mice given combination therapy compared to control mice and a corresponding decrease in pERK and cyclin D1, which are downstream targets of oncogenic Kras . Phospho-CrkL is a marker for ABL kinase activity while GAPD is a loading control. Graphs depict means and S.E.M.

    Journal: Oncotarget

    Article Title: ABL kinase inhibition sensitizes primary lung adenocarcinomas to chemotherapy by promoting tumor cell differentiation

    doi: 10.18632/oncotarget.26740

    Figure Lengend Snippet: Inhibition of ABL kinases in the presence of docetaxel decreases cell proliferation and increases cell death in Kras G12D/+ ; p53 −/− driven lung tumors Treatments began 10 weeks after Adeno-Cre infection of Rosa26-fGFP; LSL-Kras G12D/+ ; p53 fl/fl mice. Mouse lungs were harvested at 12 weeks after Adeno-Cre infection. A. IHC for Ki67+ cells in sections of mouse lungs from mice treated with vehicle control, docetaxel, GNF5, or combination (docetaxel+GNF5) treatment showing a decrease in Ki67 staining, particularly in the combination therapy group. Scale bar = 50 μm. B. Quantification of IHC staining for Ki67 ( n = 5-7 tumors per group). C. Immunofluorescence staining for Ki67 showing a decrease in the percentage of Ki67+ (red) tumor cells (labeled with farnesylated GFP, green) in mice treated with combination therapy of GNF5 and docetaxel compared to vehicle treated mice. Scale bar = 150 μm. D. Immunoblotting of lysates showed a decrease in Ki67 expression and increase in cleaved caspase 3 expression in mice given combination therapy compared to control mice and a corresponding decrease in pERK and cyclin D1, which are downstream targets of oncogenic Kras . Phospho-CrkL is a marker for ABL kinase activity while GAPD is a loading control. Graphs depict means and S.E.M.

    Article Snippet: Docetaxel (LC Laboratories: D-1000) stock solution was prepared by dissolving the powder at a concentration of 30 mg/mL in ethanol.

    Techniques: Inhibition, Infection, Mouse Assay, Immunohistochemistry, Staining, Immunofluorescence, Labeling, Expressing, Marker, Activity Assay

    Combination treatment of GNF5 and docetaxel induces cytoplasmic localization of Yap1 and decreases expression of downstream transcription targets of Yap1 compared to vehicle control treated lung adenocarcinomas GFP+ cells were isolated from KRAS G12D/+ ; p53 −/− ; Rosa26-fGFP mice two weeks after treatment with vehicle, docetaxel, GNF5, or combination treatment and 12 weeks after induction of tumors with adenovirus. A. Immunofluorescence staining for Yap1 shows a decrease in Yap1 nuclear localization in mice treated with GNF5 and docetaxel compared to vehicle control treated mice. Scale = 75 μm. B. Higher magnification images are provided to show sub-cellular localization of Yap1 in control and combination treatment mice. Scale = 7.5 μm. C. Western blot analysis showed no significant difference in total Yap1 protein expression in double treated mice compared to vehicle treated control mice. Phospho-CrkL expression is a marker of ABL kinase activity. D. RT-qPCR analysis showed a decrease in mRNA transcript expression of the downstream Yap1 target, Birc5 , which encodes the protein survivin, in mice treated with both GNF5 and docetaxel compared to vehicle control treated mice or mice treated with GNF5 or docetaxel alone ( n = 3-4 mice per group, each RT-qPCR assay performed in triplicate). E. Western blot analysis showed a decrease in protein expression of downstream transcriptional targets of Yap1, including survivin, c-Myc, S100A4, and cyclin D1. Graphs depict means and S.E.M.

    Journal: Oncotarget

    Article Title: ABL kinase inhibition sensitizes primary lung adenocarcinomas to chemotherapy by promoting tumor cell differentiation

    doi: 10.18632/oncotarget.26740

    Figure Lengend Snippet: Combination treatment of GNF5 and docetaxel induces cytoplasmic localization of Yap1 and decreases expression of downstream transcription targets of Yap1 compared to vehicle control treated lung adenocarcinomas GFP+ cells were isolated from KRAS G12D/+ ; p53 −/− ; Rosa26-fGFP mice two weeks after treatment with vehicle, docetaxel, GNF5, or combination treatment and 12 weeks after induction of tumors with adenovirus. A. Immunofluorescence staining for Yap1 shows a decrease in Yap1 nuclear localization in mice treated with GNF5 and docetaxel compared to vehicle control treated mice. Scale = 75 μm. B. Higher magnification images are provided to show sub-cellular localization of Yap1 in control and combination treatment mice. Scale = 7.5 μm. C. Western blot analysis showed no significant difference in total Yap1 protein expression in double treated mice compared to vehicle treated control mice. Phospho-CrkL expression is a marker of ABL kinase activity. D. RT-qPCR analysis showed a decrease in mRNA transcript expression of the downstream Yap1 target, Birc5 , which encodes the protein survivin, in mice treated with both GNF5 and docetaxel compared to vehicle control treated mice or mice treated with GNF5 or docetaxel alone ( n = 3-4 mice per group, each RT-qPCR assay performed in triplicate). E. Western blot analysis showed a decrease in protein expression of downstream transcriptional targets of Yap1, including survivin, c-Myc, S100A4, and cyclin D1. Graphs depict means and S.E.M.

    Article Snippet: Docetaxel (LC Laboratories: D-1000) stock solution was prepared by dissolving the powder at a concentration of 30 mg/mL in ethanol.

    Techniques: Expressing, Isolation, Mouse Assay, Immunofluorescence, Staining, Western Blot, Marker, Activity Assay, Quantitative RT-PCR

    Inhibition of ABL kinases impairs Kras G12D/+ ; p53 −/− driven lung tumors Treatments began 8 weeks after Adeno-Cre infection of LSL-Kras G12D/+ ; p53 fl/fl mice. A. 3D-reconstructions of μ-CT scans of mice before and after 14 days of treatment with vehicle, GNF5 (100 mg/kg b.i.d .), docetaxel (10 mg/kg, twice weekly), or GNF5 (100 mg/kg) + docetaxel (10 mg/kg, twice weekly). B. H E sections of lungs from mice in each treatment group 10 weeks after adenoviral induction. Scale bar = 2,000 μm. C. Quantification of total tumor volume in bilateral lungs evaluated by μ-CT scans taken before (8 weeks) and after (10 weeks) treatment with vehicle, GNF5 alone, docetaxel alone, or combination therapy shows that combination treatment significantly impairs tumor growth in mice compared to vehicle treated mice. Graphs depict means and S.E.M. of “n” mice, where “n” represents each individual animal used.

    Journal: Oncotarget

    Article Title: ABL kinase inhibition sensitizes primary lung adenocarcinomas to chemotherapy by promoting tumor cell differentiation

    doi: 10.18632/oncotarget.26740

    Figure Lengend Snippet: Inhibition of ABL kinases impairs Kras G12D/+ ; p53 −/− driven lung tumors Treatments began 8 weeks after Adeno-Cre infection of LSL-Kras G12D/+ ; p53 fl/fl mice. A. 3D-reconstructions of μ-CT scans of mice before and after 14 days of treatment with vehicle, GNF5 (100 mg/kg b.i.d .), docetaxel (10 mg/kg, twice weekly), or GNF5 (100 mg/kg) + docetaxel (10 mg/kg, twice weekly). B. H E sections of lungs from mice in each treatment group 10 weeks after adenoviral induction. Scale bar = 2,000 μm. C. Quantification of total tumor volume in bilateral lungs evaluated by μ-CT scans taken before (8 weeks) and after (10 weeks) treatment with vehicle, GNF5 alone, docetaxel alone, or combination therapy shows that combination treatment significantly impairs tumor growth in mice compared to vehicle treated mice. Graphs depict means and S.E.M. of “n” mice, where “n” represents each individual animal used.

    Article Snippet: Docetaxel (LC Laboratories: D-1000) stock solution was prepared by dissolving the powder at a concentration of 30 mg/mL in ethanol.

    Techniques: Inhibition, Infection, Mouse Assay

    Inhibition of ABL kinases sensitizes primary Kras G12D/+ derived organoid cultures to treatment with docetaxel in 3D tumor sphere assays A. Model: SPC (Sftpc)-CreERT2; KRAS LSL-G12D ; Rosa26-tdTomato mice were given tamoxifen to induce tumor formation. Tomato+ cells were then isolated from mice after tumor formation and grown in Matrigel in transwell inserts in the presence of primary mouse fibroblasts (derived from PDGFRα-H2B: GFP mice) to induce tumor organoid formation. B and C. Organoids were treated with vehicle (DMSO), GNF5, docetaxel (DOC), or combination treatment (GNF5 + DOC) for 2 weeks and assessed for organoid size 2 weeks after treatment (B); quantification revealed a significant reduction in organoid size in response to combination treatment compared to vehicle, GNF5, or docetaxel alone (C). Graphs depict means and S.E.M. D. - E. Immunofluorescence staining of organoids treated with GNF5 and docetaxel show increased expression of the Type II cell marker, SPC, compared to vehicle control treated mice. Quantification is provided in (E). Scale bar = 10 μm. Graphs depict means and S.E.M.

    Journal: Oncotarget

    Article Title: ABL kinase inhibition sensitizes primary lung adenocarcinomas to chemotherapy by promoting tumor cell differentiation

    doi: 10.18632/oncotarget.26740

    Figure Lengend Snippet: Inhibition of ABL kinases sensitizes primary Kras G12D/+ derived organoid cultures to treatment with docetaxel in 3D tumor sphere assays A. Model: SPC (Sftpc)-CreERT2; KRAS LSL-G12D ; Rosa26-tdTomato mice were given tamoxifen to induce tumor formation. Tomato+ cells were then isolated from mice after tumor formation and grown in Matrigel in transwell inserts in the presence of primary mouse fibroblasts (derived from PDGFRα-H2B: GFP mice) to induce tumor organoid formation. B and C. Organoids were treated with vehicle (DMSO), GNF5, docetaxel (DOC), or combination treatment (GNF5 + DOC) for 2 weeks and assessed for organoid size 2 weeks after treatment (B); quantification revealed a significant reduction in organoid size in response to combination treatment compared to vehicle, GNF5, or docetaxel alone (C). Graphs depict means and S.E.M. D. - E. Immunofluorescence staining of organoids treated with GNF5 and docetaxel show increased expression of the Type II cell marker, SPC, compared to vehicle control treated mice. Quantification is provided in (E). Scale bar = 10 μm. Graphs depict means and S.E.M.

    Article Snippet: Docetaxel (LC Laboratories: D-1000) stock solution was prepared by dissolving the powder at a concentration of 30 mg/mL in ethanol.

    Techniques: Inhibition, Derivative Assay, Mouse Assay, Isolation, Immunofluorescence, Staining, Expressing, Marker

    Combination treatment of GNF5 and docetaxel induces lung tumor cell differentiation in vivo GFP+ cells were isolated from KRAS LSL-G12D ; p53 fl/fl ; Rosa26-fGFP mice two weeks after treatment with vehicle, docetaxel, GNF5, or combination treatment and 12 weeks after induction of tumors with adenovirus. A. Quantification of total tumor volume in bilateral lungs evaluated by μ-CT scans taken before (10 weeks) and after (12 weeks) treatment with vehicle or combination therapy (GNF5 + docetaxel) shows that combination treatment significantly impairs tumor growth in mice compared to vehicle treated mice ( n = 11 mice per group). B. Western blot analysis showed an increase in expression of the ciliated cell marker, acetylated α-tubulin, and the secretory cell marker, Scgb1a1, with a corresponding decrease in expression of the basal cell marker, keratin 5, in mice treated with GNF5 and docetaxel. C. RT-qPCR analysis for indicated cell treatment groups shows an increase in expression of terminal cell markers ( Sftpc : Type II cell marker and Scgb1a1 : secretory cell marker) with a corresponding decrease in expression of the basal cell marker, TP63 , in mice treated with the combination therapy compared to control or mice treated with GNF5 or docetaxel alone ( n = 4-6 mice per group, each RT-PCR assay performed in triplicate). Graphs depict means and S.E.M.

    Journal: Oncotarget

    Article Title: ABL kinase inhibition sensitizes primary lung adenocarcinomas to chemotherapy by promoting tumor cell differentiation

    doi: 10.18632/oncotarget.26740

    Figure Lengend Snippet: Combination treatment of GNF5 and docetaxel induces lung tumor cell differentiation in vivo GFP+ cells were isolated from KRAS LSL-G12D ; p53 fl/fl ; Rosa26-fGFP mice two weeks after treatment with vehicle, docetaxel, GNF5, or combination treatment and 12 weeks after induction of tumors with adenovirus. A. Quantification of total tumor volume in bilateral lungs evaluated by μ-CT scans taken before (10 weeks) and after (12 weeks) treatment with vehicle or combination therapy (GNF5 + docetaxel) shows that combination treatment significantly impairs tumor growth in mice compared to vehicle treated mice ( n = 11 mice per group). B. Western blot analysis showed an increase in expression of the ciliated cell marker, acetylated α-tubulin, and the secretory cell marker, Scgb1a1, with a corresponding decrease in expression of the basal cell marker, keratin 5, in mice treated with GNF5 and docetaxel. C. RT-qPCR analysis for indicated cell treatment groups shows an increase in expression of terminal cell markers ( Sftpc : Type II cell marker and Scgb1a1 : secretory cell marker) with a corresponding decrease in expression of the basal cell marker, TP63 , in mice treated with the combination therapy compared to control or mice treated with GNF5 or docetaxel alone ( n = 4-6 mice per group, each RT-PCR assay performed in triplicate). Graphs depict means and S.E.M.

    Article Snippet: Docetaxel (LC Laboratories: D-1000) stock solution was prepared by dissolving the powder at a concentration of 30 mg/mL in ethanol.

    Techniques: Cell Differentiation, In Vivo, Isolation, Mouse Assay, Western Blot, Expressing, Marker, Quantitative RT-PCR, Reverse Transcription Polymerase Chain Reaction

    Forkhead box protein M1 (FOXM1) promotes docetaxel resistance through regulating the microtubule-destabilizing protein Stathmin rather than mitotic centromere–associated kinesin (MCAK) in gastric cancers. ( A ) mRNA levels of FOXM1, Stathmin and MCAK in AGS cells after transfected with pcDNA3. 1, pcDNA3. 1-FOXM1, non-specific siRNA or FOXM1-siRNA were determined by RT-PCR. ( B ) The protein expression levels of FOXM1, Stathmin and MCAK after transfections were shown by western blot analysis. ( C ) Polymerized and soluble tubulin fractions from siRNA -FOXM1, siRNA-Stathmin, siRNA-MCAK and non-specific siRNA-transfected AGS-DOC R cell lines treated or untreated with docetaxel were detected by western blot (Left). The relative polymerized microtubule fractions of both α-tubulin and β-tubulin were significantly increased after the treatment of docetaxel in FOXM1, Stathmin and MCAK knockdown AGS-DOC R cells (Right). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin

    doi: 10.1111/jcmm.12216

    Figure Lengend Snippet: Forkhead box protein M1 (FOXM1) promotes docetaxel resistance through regulating the microtubule-destabilizing protein Stathmin rather than mitotic centromere–associated kinesin (MCAK) in gastric cancers. ( A ) mRNA levels of FOXM1, Stathmin and MCAK in AGS cells after transfected with pcDNA3. 1, pcDNA3. 1-FOXM1, non-specific siRNA or FOXM1-siRNA were determined by RT-PCR. ( B ) The protein expression levels of FOXM1, Stathmin and MCAK after transfections were shown by western blot analysis. ( C ) Polymerized and soluble tubulin fractions from siRNA -FOXM1, siRNA-Stathmin, siRNA-MCAK and non-specific siRNA-transfected AGS-DOC R cell lines treated or untreated with docetaxel were detected by western blot (Left). The relative polymerized microtubule fractions of both α-tubulin and β-tubulin were significantly increased after the treatment of docetaxel in FOXM1, Stathmin and MCAK knockdown AGS-DOC R cells (Right). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Article Snippet: The semisynthetic taxane, docetaxel (Selleckchem, Houston, TX, USA), was dissolved in dimethyl sulfoxide (DMSO) and diluted to a final concentration of 0.015, 0.020 and 0.15 mg/l, whereas thiazole antibiotic thiostrepton (BBI, Boston, MA, USA), which has been previously shown to inhibit FOXM1 expression, was prepared to be a solution at the concentration of 16 mg/l with DMSO before use [ ].

    Techniques: Transfection, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot

    Thiostrepton can reverse docetaxel resistance in gastric cancer cells. AGS-DOC R cells were treated with dimethyl sulfoxide (vehicle control), 0.15 mg/l docetaxel, 16 mg/l thiostrepton or a combination of 0.15 mg/l docetaxel and 16 mg/l thiostrepton for 72 hrs. ( A ) The percentage of viable cells in different treatment group is shown at each time-point by MTT assay. ( B ) Cell lysates were prepared at 0, 24, 48 and 72 hrs after treatment, and the expression of forkhead box protein M1, Stathmin and GAPDH were analysed by western blot analysis.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin

    doi: 10.1111/jcmm.12216

    Figure Lengend Snippet: Thiostrepton can reverse docetaxel resistance in gastric cancer cells. AGS-DOC R cells were treated with dimethyl sulfoxide (vehicle control), 0.15 mg/l docetaxel, 16 mg/l thiostrepton or a combination of 0.15 mg/l docetaxel and 16 mg/l thiostrepton for 72 hrs. ( A ) The percentage of viable cells in different treatment group is shown at each time-point by MTT assay. ( B ) Cell lysates were prepared at 0, 24, 48 and 72 hrs after treatment, and the expression of forkhead box protein M1, Stathmin and GAPDH were analysed by western blot analysis.

    Article Snippet: The semisynthetic taxane, docetaxel (Selleckchem, Houston, TX, USA), was dissolved in dimethyl sulfoxide (DMSO) and diluted to a final concentration of 0.015, 0.020 and 0.15 mg/l, whereas thiazole antibiotic thiostrepton (BBI, Boston, MA, USA), which has been previously shown to inhibit FOXM1 expression, was prepared to be a solution at the concentration of 16 mg/l with DMSO before use [ ].

    Techniques: MTT Assay, Expressing, Western Blot

    Forkhead box protein M1 (FOXM1) correlated with Stathmin in gastric cancers. ( A ) The luciferase activity of Stathmin promoter–reporter vectors was modulated by FOXM1 levels in human gastric cell lines. (Left) MNK-28 and AGS cells were transfected with Stathmin promoter–reporter vectors and increasing amounts (0.2, 0.4, 0.6 ng) of pcDNA3, 1-FOXM1. (Right) AGS and SGC-7901 cells were transfected with Stathmin promoter–reporter vectors and transfection reagents, non-specific siRNA or different amount (10 and 20 nM) of FOXM1-siRNA. pcDNA3,1 was used as a control. The relative Stathmin promoter activities were measured 24 hrs after transfection, and the activities in the treated groups were expressed as the fold or percentage of that in their respective control groups. ( B ) Chromatin immunoprecipitation assay (ChIP) was performed in AGS and MKN-28 cells using an antibody specific to FOXM1 or IgG as a control. PCR was used to amplify the region surrounding the putative FOXM1 binding site at −5793 upstream of the transcriptional start site and the region surrounding −1251 as a non-specific control. Representative PCR results are shown. ( C ) Silencing FOXM1 sensitized docetaxel-resistant gastric cancer cells, and Stathmin overexpression rescued chemoresistance in FOXM1-silenced AGS-DOC R cells. Western blot analysis reveals the expression of genes in AGS-DOC R cells expressing pcDNA3,1-Stathmin, si-FOXM1, si-Stathmin or si-FOXM1 plus Stathmin (Top). MTT assay shows cell survival in AGS-DOC R cells expressing pcDNA3,1, pcDNA3,1-Stathmin, non-specific siRNA, si-FOXM1, si-Stathmin or si-FOXM1 plus Stathmin treated with docetaxel and the IC50 was 0.026, 0.036, 0.024, 0.011, 0.014, 0.019 mg/l respectively (Bottom). ( D ) Immunohistochemical staining for FOXM1 and Stathmin antibody in human gastric cancer tissues (×400). The paraffin-embedded gastric tissues were stained with antibodies FOXM1 (A, B, C, D) and Stathmin (E, F, G, H). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significantly.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin

    doi: 10.1111/jcmm.12216

    Figure Lengend Snippet: Forkhead box protein M1 (FOXM1) correlated with Stathmin in gastric cancers. ( A ) The luciferase activity of Stathmin promoter–reporter vectors was modulated by FOXM1 levels in human gastric cell lines. (Left) MNK-28 and AGS cells were transfected with Stathmin promoter–reporter vectors and increasing amounts (0.2, 0.4, 0.6 ng) of pcDNA3, 1-FOXM1. (Right) AGS and SGC-7901 cells were transfected with Stathmin promoter–reporter vectors and transfection reagents, non-specific siRNA or different amount (10 and 20 nM) of FOXM1-siRNA. pcDNA3,1 was used as a control. The relative Stathmin promoter activities were measured 24 hrs after transfection, and the activities in the treated groups were expressed as the fold or percentage of that in their respective control groups. ( B ) Chromatin immunoprecipitation assay (ChIP) was performed in AGS and MKN-28 cells using an antibody specific to FOXM1 or IgG as a control. PCR was used to amplify the region surrounding the putative FOXM1 binding site at −5793 upstream of the transcriptional start site and the region surrounding −1251 as a non-specific control. Representative PCR results are shown. ( C ) Silencing FOXM1 sensitized docetaxel-resistant gastric cancer cells, and Stathmin overexpression rescued chemoresistance in FOXM1-silenced AGS-DOC R cells. Western blot analysis reveals the expression of genes in AGS-DOC R cells expressing pcDNA3,1-Stathmin, si-FOXM1, si-Stathmin or si-FOXM1 plus Stathmin (Top). MTT assay shows cell survival in AGS-DOC R cells expressing pcDNA3,1, pcDNA3,1-Stathmin, non-specific siRNA, si-FOXM1, si-Stathmin or si-FOXM1 plus Stathmin treated with docetaxel and the IC50 was 0.026, 0.036, 0.024, 0.011, 0.014, 0.019 mg/l respectively (Bottom). ( D ) Immunohistochemical staining for FOXM1 and Stathmin antibody in human gastric cancer tissues (×400). The paraffin-embedded gastric tissues were stained with antibodies FOXM1 (A, B, C, D) and Stathmin (E, F, G, H). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significantly.

    Article Snippet: The semisynthetic taxane, docetaxel (Selleckchem, Houston, TX, USA), was dissolved in dimethyl sulfoxide (DMSO) and diluted to a final concentration of 0.015, 0.020 and 0.15 mg/l, whereas thiazole antibiotic thiostrepton (BBI, Boston, MA, USA), which has been previously shown to inhibit FOXM1 expression, was prepared to be a solution at the concentration of 16 mg/l with DMSO before use [ ].

    Techniques: Luciferase, Activity Assay, Transfection, Chromatin Immunoprecipitation, Polymerase Chain Reaction, Binding Assay, Over Expression, Western Blot, Expressing, MTT Assay, Immunohistochemistry, Staining

    Forkhead box protein M1 (FOXM1) alters the microtubule dynamics in preventing docetaxel-induced apoptosis. Polymerized and soluble tubulin fractions from docetaxel untreated and treated FOXM1-siRNA and pcDNA3, 1-FOXM1–transfected cell lines were generated by centrifugation. Western blot was used to assay α-tubulin and β-tubulin ratios in polymerized and soluble fractions. Relative percentages are shown above western blot (Left). The soluble-to-polymerized microtubule fractions after docetaxel treatment were significantly inhibited in FOXM1-overexpressed group, analysed by t -test both for α-tubulin and for β-tubulin (Right). ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin

    doi: 10.1111/jcmm.12216

    Figure Lengend Snippet: Forkhead box protein M1 (FOXM1) alters the microtubule dynamics in preventing docetaxel-induced apoptosis. Polymerized and soluble tubulin fractions from docetaxel untreated and treated FOXM1-siRNA and pcDNA3, 1-FOXM1–transfected cell lines were generated by centrifugation. Western blot was used to assay α-tubulin and β-tubulin ratios in polymerized and soluble fractions. Relative percentages are shown above western blot (Left). The soluble-to-polymerized microtubule fractions after docetaxel treatment were significantly inhibited in FOXM1-overexpressed group, analysed by t -test both for α-tubulin and for β-tubulin (Right). ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Article Snippet: The semisynthetic taxane, docetaxel (Selleckchem, Houston, TX, USA), was dissolved in dimethyl sulfoxide (DMSO) and diluted to a final concentration of 0.015, 0.020 and 0.15 mg/l, whereas thiazole antibiotic thiostrepton (BBI, Boston, MA, USA), which has been previously shown to inhibit FOXM1 expression, was prepared to be a solution at the concentration of 16 mg/l with DMSO before use [ ].

    Techniques: Transfection, Generated, Centrifugation, Western Blot

    Elevated levels of forkhead box protein M1 (FOXM1) correlate with resistance to docetaxel in gastric cancer. ( A , Top) The expression of FOXM1 in three gastric cancer cell lines: AGS, SGC-7901 and MKN-28, shown by western blot. (Bottom) The expression of FOXM1 after transfected with pcDNA3. 1, pcDNA3. 1-FOXM1, non-specific siRNA or FOXM1-siRNA in gastric cell lines AGS, analysed by western blot 48 hrs later. ( B ) AGS, SGC-7901 and MKN-28 cells were treated with 0.02 mg/l of docetaxel for 0, 24, 48 and 72 hrs. MTT assay was performed to test the cell viability. ( C ) Gastric cell lines AGS were treated with docetaxel at the concentration of 0.02 mg/l after FOXM1-siRNA or pcDNA3, 1-FOXM1 transfection for 72 hrs. Cell growth curves were drawn by MTT assays. The IC50 in FOXM1 knockdown, overexpressed, non-specific siRNA and pcDNA3, 1 transfected groups was 0.012, 0.040, 0.024 and 0.027 mg/l respectively ( D ). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin

    doi: 10.1111/jcmm.12216

    Figure Lengend Snippet: Elevated levels of forkhead box protein M1 (FOXM1) correlate with resistance to docetaxel in gastric cancer. ( A , Top) The expression of FOXM1 in three gastric cancer cell lines: AGS, SGC-7901 and MKN-28, shown by western blot. (Bottom) The expression of FOXM1 after transfected with pcDNA3. 1, pcDNA3. 1-FOXM1, non-specific siRNA or FOXM1-siRNA in gastric cell lines AGS, analysed by western blot 48 hrs later. ( B ) AGS, SGC-7901 and MKN-28 cells were treated with 0.02 mg/l of docetaxel for 0, 24, 48 and 72 hrs. MTT assay was performed to test the cell viability. ( C ) Gastric cell lines AGS were treated with docetaxel at the concentration of 0.02 mg/l after FOXM1-siRNA or pcDNA3, 1-FOXM1 transfection for 72 hrs. Cell growth curves were drawn by MTT assays. The IC50 in FOXM1 knockdown, overexpressed, non-specific siRNA and pcDNA3, 1 transfected groups was 0.012, 0.040, 0.024 and 0.027 mg/l respectively ( D ). * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Article Snippet: The semisynthetic taxane, docetaxel (Selleckchem, Houston, TX, USA), was dissolved in dimethyl sulfoxide (DMSO) and diluted to a final concentration of 0.015, 0.020 and 0.15 mg/l, whereas thiazole antibiotic thiostrepton (BBI, Boston, MA, USA), which has been previously shown to inhibit FOXM1 expression, was prepared to be a solution at the concentration of 16 mg/l with DMSO before use [ ].

    Techniques: Expressing, Western Blot, Transfection, MTT Assay, Concentration Assay

    Docetaxel-resistant cell line shows elevated forkhead box protein M1 (FOXM1) mRNA and protein expression levels. ( A ) Cells after molecular evolution assay were treated with increasing concentrations of docetaxel, respectively, and their rates of cell viability were measured by MTT assay. R0 represents the untreated control cell line, whereas R2, R4, R6 and R8 represent AGS cells that are treated with 0.015 mg/l docetaxel for two, four, six and eight times, separately. IC50 for cells in R0, R2, R4, R6 and R8 was 0.026, 0.033, 0.054, 0.098, 0.190 mg/l. ( B ) FOXM1 mRNA transcript levels in R0, R2, R4, R6 and R8 cells were determined by RT-PCR analysis. ( C ) Western blot analysis was performed to detect the relative protein expression levels of FOXM1, Stathmin and mitotic centromere–associated kinesin in the different treated round. Results shown were derived from at least three independent experiments. Statistical analysis was performed with Student*s t -tests. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Journal: Journal of Cellular and Molecular Medicine

    Article Title: FOXM1 mediates resistance to docetaxel in gastric cancer via up-regulating Stathmin

    doi: 10.1111/jcmm.12216

    Figure Lengend Snippet: Docetaxel-resistant cell line shows elevated forkhead box protein M1 (FOXM1) mRNA and protein expression levels. ( A ) Cells after molecular evolution assay were treated with increasing concentrations of docetaxel, respectively, and their rates of cell viability were measured by MTT assay. R0 represents the untreated control cell line, whereas R2, R4, R6 and R8 represent AGS cells that are treated with 0.015 mg/l docetaxel for two, four, six and eight times, separately. IC50 for cells in R0, R2, R4, R6 and R8 was 0.026, 0.033, 0.054, 0.098, 0.190 mg/l. ( B ) FOXM1 mRNA transcript levels in R0, R2, R4, R6 and R8 cells were determined by RT-PCR analysis. ( C ) Western blot analysis was performed to detect the relative protein expression levels of FOXM1, Stathmin and mitotic centromere–associated kinesin in the different treated round. Results shown were derived from at least three independent experiments. Statistical analysis was performed with Student*s t -tests. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001 significant.

    Article Snippet: The semisynthetic taxane, docetaxel (Selleckchem, Houston, TX, USA), was dissolved in dimethyl sulfoxide (DMSO) and diluted to a final concentration of 0.015, 0.020 and 0.15 mg/l, whereas thiazole antibiotic thiostrepton (BBI, Boston, MA, USA), which has been previously shown to inhibit FOXM1 expression, was prepared to be a solution at the concentration of 16 mg/l with DMSO before use [ ].

    Techniques: Expressing, MTT Assay, Reverse Transcription Polymerase Chain Reaction, Western Blot, Derivative Assay

    Docetaxel induces enlargement and expansion of tumor-draining lymphatics in vivo that can be attenuated by VEGFR3 blockade. a Quantified lymphatic vessel perimeter in both tumor-bearing mammary fat pad of treated 4T1 mice and contralateral naïve (non-tumor bearing) fat pad. b Quantified lymphatic vessel area in both tumor-bearing mammary fat pad of treated 4T1 mice and contralateral naïve (non-tumor bearing) fat pad. c Quantified lymphatic vessel density of single podoplanin+ lymphatic vessels per stromal area throughout sections. d Total lymphatic endothelial cells per weight of fat pad (live CD45 − CD31 + gp38 + ) as determined by flow cytometry. e Fluid drainage from tumor to axillary lymph node as determined after Evans blue injection into tumor-bearing mammary fat pad 24 h after treatment with docetaxel. ( n = 5–9 mice per group). f Quantified lymphatic vessel density displayed as number of peritumoral lymphatic vessels per mm 2 of stromal tissue after 0, 1, 2, and 3 doses of docetaxel (8 mg/kg, IV, 3 days apart). * p

    Journal: BMC Cancer

    Article Title: Docetaxel facilitates lymphatic-tumor crosstalk to promote lymphangiogenesis and cancer progression

    doi: 10.1186/s12885-018-4619-8

    Figure Lengend Snippet: Docetaxel induces enlargement and expansion of tumor-draining lymphatics in vivo that can be attenuated by VEGFR3 blockade. a Quantified lymphatic vessel perimeter in both tumor-bearing mammary fat pad of treated 4T1 mice and contralateral naïve (non-tumor bearing) fat pad. b Quantified lymphatic vessel area in both tumor-bearing mammary fat pad of treated 4T1 mice and contralateral naïve (non-tumor bearing) fat pad. c Quantified lymphatic vessel density of single podoplanin+ lymphatic vessels per stromal area throughout sections. d Total lymphatic endothelial cells per weight of fat pad (live CD45 − CD31 + gp38 + ) as determined by flow cytometry. e Fluid drainage from tumor to axillary lymph node as determined after Evans blue injection into tumor-bearing mammary fat pad 24 h after treatment with docetaxel. ( n = 5–9 mice per group). f Quantified lymphatic vessel density displayed as number of peritumoral lymphatic vessels per mm 2 of stromal tissue after 0, 1, 2, and 3 doses of docetaxel (8 mg/kg, IV, 3 days apart). * p

    Article Snippet: Once tumors were palpable (5 days post-inoculation), mice underwent three consecutive days of treatment beginning the following day consisting of anti-VEGFR3 antibody (200 μg, I.P., eBioscience) or IgG control on the first day, followed by a single dose of docetaxel in saline/20% ethanol (8 mg/kg I.V., Enzo Life Sciences) or saline/20% ethanol control on the second day, and another treatment with anti-VEGFR3 antibody or IgG control on the third day.

    Techniques: In Vivo, Mouse Assay, Flow Cytometry, Cytometry, Injection

    Docetaxel treatment increases expression of pro-lymphangiogenic cytokines in 4T1 tumors. a Heat map representation of expression of chemokines associated with lymphangiogenesis (left) and corresponding known roles in cancer and lymphangiogenesis (right). Results obtained by flow cytometry analysis of 4T1 tumors treated as outlined in Fig. 2 ( n = 4/group). Log-transformed data displayed as fold change and heat map generated using MatLab software. (*) indicates analysis by ELISA. Docetaxel abbreviated as DTX and anti-VEGFR3 therapy abbreviated as α-V3 in figure. b Proposed mechanism by which docetaxel results in lymphatic activation and the resulting effect on cancer cell response to therapy. (1) Docetaxel induces production of pro-lymphangiogenic factors in the breast tumor microenvironment. (2) Docetaxel-induced lymphangiogenic factors like VEGFC and TNFa result in VEGFR3-dependent enlargement of lymphatics (brown) and lymphangiogenesis. (3) Docetaxel-activated lymphatics promote VEGFR3-mediated tumor cell (green) invasion and (4) reduce docetaxel efficacy

    Journal: BMC Cancer

    Article Title: Docetaxel facilitates lymphatic-tumor crosstalk to promote lymphangiogenesis and cancer progression

    doi: 10.1186/s12885-018-4619-8

    Figure Lengend Snippet: Docetaxel treatment increases expression of pro-lymphangiogenic cytokines in 4T1 tumors. a Heat map representation of expression of chemokines associated with lymphangiogenesis (left) and corresponding known roles in cancer and lymphangiogenesis (right). Results obtained by flow cytometry analysis of 4T1 tumors treated as outlined in Fig. 2 ( n = 4/group). Log-transformed data displayed as fold change and heat map generated using MatLab software. (*) indicates analysis by ELISA. Docetaxel abbreviated as DTX and anti-VEGFR3 therapy abbreviated as α-V3 in figure. b Proposed mechanism by which docetaxel results in lymphatic activation and the resulting effect on cancer cell response to therapy. (1) Docetaxel induces production of pro-lymphangiogenic factors in the breast tumor microenvironment. (2) Docetaxel-induced lymphangiogenic factors like VEGFC and TNFa result in VEGFR3-dependent enlargement of lymphatics (brown) and lymphangiogenesis. (3) Docetaxel-activated lymphatics promote VEGFR3-mediated tumor cell (green) invasion and (4) reduce docetaxel efficacy

    Article Snippet: Once tumors were palpable (5 days post-inoculation), mice underwent three consecutive days of treatment beginning the following day consisting of anti-VEGFR3 antibody (200 μg, I.P., eBioscience) or IgG control on the first day, followed by a single dose of docetaxel in saline/20% ethanol (8 mg/kg I.V., Enzo Life Sciences) or saline/20% ethanol control on the second day, and another treatment with anti-VEGFR3 antibody or IgG control on the third day.

    Techniques: Expressing, Flow Cytometry, Cytometry, Transformation Assay, Generated, Software, Enzyme-linked Immunosorbent Assay, Activation Assay

    Docetaxel induces invasion of multiple human breast cancer cell lines toward lymphatics in vitro in a VEGFR3-dependent manner. a Schematic of the in vitro tissue engineered model of the tumor-lymphatic interface in the human breast cancer microenvironment. Our model contains mammary stromal fibroblasts and TNBC cells in a collagen I matrix. LECs are seeded along the underside of the insert system through which tumor cells transmigrate from a basal to luminal fashion. Physiologically relevant flow (1 μm/s) is applied via a pressure head of media to yield delivery of docetaxel. Schematic depicts experimental groups. b Fold change in invasion of MDA-MB-231 tumor cells across the porous membrane in our 3D microenvironment system +/− docetaxel treatment (0.1 μM), +/− MAZ51 (1 μM), and/or in the presence or absence of LECs. c Fold change in invasion of HCC38 tumor cells across the porous membrane in our 3D microenvironment system +/− docetaxel treatment (1 μM), +/− MAZ51 (1 μM), and/or in the presence or absence of LECs. d Fold change in invasion of HCC1806 tumor cells across the porous membrane in our 3D microenvironment system +/− docetaxel treatment (0.1 μM), +/− MAZ51 (1 μM), and/or in the presence or absence of LECs. Fold change calculated as compared to no docetaxel/with LEC control. n ≥ 3 biological replicates. * p

    Journal: BMC Cancer

    Article Title: Docetaxel facilitates lymphatic-tumor crosstalk to promote lymphangiogenesis and cancer progression

    doi: 10.1186/s12885-018-4619-8

    Figure Lengend Snippet: Docetaxel induces invasion of multiple human breast cancer cell lines toward lymphatics in vitro in a VEGFR3-dependent manner. a Schematic of the in vitro tissue engineered model of the tumor-lymphatic interface in the human breast cancer microenvironment. Our model contains mammary stromal fibroblasts and TNBC cells in a collagen I matrix. LECs are seeded along the underside of the insert system through which tumor cells transmigrate from a basal to luminal fashion. Physiologically relevant flow (1 μm/s) is applied via a pressure head of media to yield delivery of docetaxel. Schematic depicts experimental groups. b Fold change in invasion of MDA-MB-231 tumor cells across the porous membrane in our 3D microenvironment system +/− docetaxel treatment (0.1 μM), +/− MAZ51 (1 μM), and/or in the presence or absence of LECs. c Fold change in invasion of HCC38 tumor cells across the porous membrane in our 3D microenvironment system +/− docetaxel treatment (1 μM), +/− MAZ51 (1 μM), and/or in the presence or absence of LECs. d Fold change in invasion of HCC1806 tumor cells across the porous membrane in our 3D microenvironment system +/− docetaxel treatment (0.1 μM), +/− MAZ51 (1 μM), and/or in the presence or absence of LECs. Fold change calculated as compared to no docetaxel/with LEC control. n ≥ 3 biological replicates. * p

    Article Snippet: Once tumors were palpable (5 days post-inoculation), mice underwent three consecutive days of treatment beginning the following day consisting of anti-VEGFR3 antibody (200 μg, I.P., eBioscience) or IgG control on the first day, followed by a single dose of docetaxel in saline/20% ethanol (8 mg/kg I.V., Enzo Life Sciences) or saline/20% ethanol control on the second day, and another treatment with anti-VEGFR3 antibody or IgG control on the third day.

    Techniques: In Vitro, Flow Cytometry, Multiple Displacement Amplification

    Blockade of VEGFR3 synergizes with docetaxel to reduce tumor growth and docetaxel-enhanced metastasis in 4T1 breast cancer. a Top panel , Representative images of 4T1 breast tumor cells (red) in the inguinal lymph nodes of mice treated with systemic docetaxel, 8 mg/kg IV (or vehicle control) and/or anti-VEGFR3 antibody, 400 μg total over 2 doses, IP (or control IgG) as detected by histological analysis of RFP-expressing tumor cells. Scale bar = 500 μm. Bottom panel, magnified images from boxed regions in top panel. Dotted white lines outline lymph node border. Scale bar = 100 μm. b Quantification of lymph node metastasis from whole lymph node scans as percent coverage of RFP+ area in whole lymph node sections. ( n ≥ 4/group) ( c ) Tumor volume of treated mice (mm 3 ) of 4T1 mice treated as described above via caliper measurements. Blue dashed arrow indicates dosing of anti-VEGFR3 antibody or control IgG and red dashed arrow indicates dosing of docetaxel or vehicle control. Curve was analyzed by MANOVA ( n = 5/group) with * p

    Journal: BMC Cancer

    Article Title: Docetaxel facilitates lymphatic-tumor crosstalk to promote lymphangiogenesis and cancer progression

    doi: 10.1186/s12885-018-4619-8

    Figure Lengend Snippet: Blockade of VEGFR3 synergizes with docetaxel to reduce tumor growth and docetaxel-enhanced metastasis in 4T1 breast cancer. a Top panel , Representative images of 4T1 breast tumor cells (red) in the inguinal lymph nodes of mice treated with systemic docetaxel, 8 mg/kg IV (or vehicle control) and/or anti-VEGFR3 antibody, 400 μg total over 2 doses, IP (or control IgG) as detected by histological analysis of RFP-expressing tumor cells. Scale bar = 500 μm. Bottom panel, magnified images from boxed regions in top panel. Dotted white lines outline lymph node border. Scale bar = 100 μm. b Quantification of lymph node metastasis from whole lymph node scans as percent coverage of RFP+ area in whole lymph node sections. ( n ≥ 4/group) ( c ) Tumor volume of treated mice (mm 3 ) of 4T1 mice treated as described above via caliper measurements. Blue dashed arrow indicates dosing of anti-VEGFR3 antibody or control IgG and red dashed arrow indicates dosing of docetaxel or vehicle control. Curve was analyzed by MANOVA ( n = 5/group) with * p

    Article Snippet: Once tumors were palpable (5 days post-inoculation), mice underwent three consecutive days of treatment beginning the following day consisting of anti-VEGFR3 antibody (200 μg, I.P., eBioscience) or IgG control on the first day, followed by a single dose of docetaxel in saline/20% ethanol (8 mg/kg I.V., Enzo Life Sciences) or saline/20% ethanol control on the second day, and another treatment with anti-VEGFR3 antibody or IgG control on the third day.

    Techniques: Mouse Assay, Expressing

    Docetaxel induces VEGFR3-dependent morphological changes of tumor-draining lymphatics in vivo. Representative images of 4T1 tumor-bearing fat pad tissue sections from mice treated as outlined in Fig. 1 . Sections were immunostained for lymphatic marker podoplanin (black) and nuclei (green). Left panel (scale bar = 4 mm) shows whole tissue sections and middle panel (scale bar = 500 μm) shows peritumoral stroma. Red dashed lines show border of inguinal lymph node, blue dashed line shows border of tumor, and black arrows indicate lymphatic vessels. Right panel (scale bar = 125 μm) shows representative images of morphology of individual lymphatic vessels

    Journal: BMC Cancer

    Article Title: Docetaxel facilitates lymphatic-tumor crosstalk to promote lymphangiogenesis and cancer progression

    doi: 10.1186/s12885-018-4619-8

    Figure Lengend Snippet: Docetaxel induces VEGFR3-dependent morphological changes of tumor-draining lymphatics in vivo. Representative images of 4T1 tumor-bearing fat pad tissue sections from mice treated as outlined in Fig. 1 . Sections were immunostained for lymphatic marker podoplanin (black) and nuclei (green). Left panel (scale bar = 4 mm) shows whole tissue sections and middle panel (scale bar = 500 μm) shows peritumoral stroma. Red dashed lines show border of inguinal lymph node, blue dashed line shows border of tumor, and black arrows indicate lymphatic vessels. Right panel (scale bar = 125 μm) shows representative images of morphology of individual lymphatic vessels

    Article Snippet: Once tumors were palpable (5 days post-inoculation), mice underwent three consecutive days of treatment beginning the following day consisting of anti-VEGFR3 antibody (200 μg, I.P., eBioscience) or IgG control on the first day, followed by a single dose of docetaxel in saline/20% ethanol (8 mg/kg I.V., Enzo Life Sciences) or saline/20% ethanol control on the second day, and another treatment with anti-VEGFR3 antibody or IgG control on the third day.

    Techniques: In Vivo, Mouse Assay, Marker

    Lymphatic endothelial cells diminish cancer cell response to docetaxel. a Cancer cell death (% dead cells of total) of MDA-MB-231 cells in 3D microenvironment system +/− docetaxel treatment (10 μM) +/− LECs as measured by live/dead stain in flow cytometry. b Cancer cell death (% dead cells of total) of HCC38 cells in 3D microenvironment system +/- docetaxel treatment (10 μM) +/− LECs. c Cancer cell death (% dead cells of total) of HCC1806 cells in 3D microenvironment system +/- docetaxel treatment (10 μM) +/− LECs. d LEC-conditioned media (LEC CM) was administered to MDA-MB-231, HCC38, or HCC1806 cells followed by 1 μM docetaxel and viability assessed by CCK8 analysis. Results displayed as measured absorbance. * p

    Journal: BMC Cancer

    Article Title: Docetaxel facilitates lymphatic-tumor crosstalk to promote lymphangiogenesis and cancer progression

    doi: 10.1186/s12885-018-4619-8

    Figure Lengend Snippet: Lymphatic endothelial cells diminish cancer cell response to docetaxel. a Cancer cell death (% dead cells of total) of MDA-MB-231 cells in 3D microenvironment system +/− docetaxel treatment (10 μM) +/− LECs as measured by live/dead stain in flow cytometry. b Cancer cell death (% dead cells of total) of HCC38 cells in 3D microenvironment system +/- docetaxel treatment (10 μM) +/− LECs. c Cancer cell death (% dead cells of total) of HCC1806 cells in 3D microenvironment system +/- docetaxel treatment (10 μM) +/− LECs. d LEC-conditioned media (LEC CM) was administered to MDA-MB-231, HCC38, or HCC1806 cells followed by 1 μM docetaxel and viability assessed by CCK8 analysis. Results displayed as measured absorbance. * p

    Article Snippet: Once tumors were palpable (5 days post-inoculation), mice underwent three consecutive days of treatment beginning the following day consisting of anti-VEGFR3 antibody (200 μg, I.P., eBioscience) or IgG control on the first day, followed by a single dose of docetaxel in saline/20% ethanol (8 mg/kg I.V., Enzo Life Sciences) or saline/20% ethanol control on the second day, and another treatment with anti-VEGFR3 antibody or IgG control on the third day.

    Techniques: Multiple Displacement Amplification, Staining, Flow Cytometry, Cytometry

    Effects of docetaxel on ( A ) SKOV3 and SKOV3-TR or ( B ) HeyA8 and HeyA8-MDR ovarian cancer cells. The percentage of apoptosis was determined by terminal deoxynucleotidyl transferase – mediated nick end labeling. Cells were treated with or without

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effects of docetaxel on ( A ) SKOV3 and SKOV3-TR or ( B ) HeyA8 and HeyA8-MDR ovarian cancer cells. The percentage of apoptosis was determined by terminal deoxynucleotidyl transferase – mediated nick end labeling. Cells were treated with or without

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques: End Labeling

    Effect of docetaxel on ovarian cancer cell growth: ( A ) HeyA8 and HeyA8-MDR or ( B ) SKOV3 or SKOV3-TR cells were plated in 96-well plates and subsequently incubated with increasing concentrations of docetaxel for 96 hours, and cell viability was determined.

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effect of docetaxel on ovarian cancer cell growth: ( A ) HeyA8 and HeyA8-MDR or ( B ) SKOV3 or SKOV3-TR cells were plated in 96-well plates and subsequently incubated with increasing concentrations of docetaxel for 96 hours, and cell viability was determined.

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques: Incubation

    Effect of docetaxel on caspase-3 ( A ), caspase-8 ( B ), and caspase-9 ( C ) activity. Ovarian cancer cells were treated with docetaxel for 24 hours followed by fluorometric profiling of caspase activity using a commercially available kit. Columns, means of

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effect of docetaxel on caspase-3 ( A ), caspase-8 ( B ), and caspase-9 ( C ) activity. Ovarian cancer cells were treated with docetaxel for 24 hours followed by fluorometric profiling of caspase activity using a commercially available kit. Columns, means of

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques: Activity Assay

    Docetaxel treatment results in FAK cleavage. The taxane-sensitive ( A ) and taxane-resistant ( B ) SKOV3 cells were cultured in the presence of docetaxel for 48 or 72 hours. Western blot analysis for FAK in SKOV3 and SKOV3-TR cells in the presence or absence

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Docetaxel treatment results in FAK cleavage. The taxane-sensitive ( A ) and taxane-resistant ( B ) SKOV3 cells were cultured in the presence of docetaxel for 48 or 72 hours. Western blot analysis for FAK in SKOV3 and SKOV3-TR cells in the presence or absence

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques: Cell Culture, Western Blot

    Effect of FAK silencing on docetaxel-sensitivity in ovarian cancer cell lines. A, Western blot analysis of SKOV3 whole-cell lysates probed with anti-FAK and anti-actin monoclonal antibodies. Bottom, densitometry results. Control siRNA did not significantly

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Effect of FAK silencing on docetaxel-sensitivity in ovarian cancer cell lines. A, Western blot analysis of SKOV3 whole-cell lysates probed with anti-FAK and anti-actin monoclonal antibodies. Bottom, densitometry results. Control siRNA did not significantly

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques: Western Blot

    Docetaxel-mediated apoptosis with or without FAK siRNA in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR ovarian cancer cells. Columns, means of three independent experiments; bars, SE. Doc, docetaxel.

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: Docetaxel-mediated apoptosis with or without FAK siRNA in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR ovarian cancer cells. Columns, means of three independent experiments; bars, SE. Doc, docetaxel.

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques:

    FAK gene silencing potentiates docetaxel-induced caspase-3 activity in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR cells. Columns, means of three experiments; bars, SE. *, P

    Journal:

    Article Title: Focal Adhesion Kinase Silencing Augments Docetaxel-Mediated Apoptosis in Ovarian Cancer Cells

    doi: 10.1158/1078-0432.CCR-05-1728

    Figure Lengend Snippet: FAK gene silencing potentiates docetaxel-induced caspase-3 activity in ( A ) SKOV3 and HeyA8 and ( B ) SKOV3-TR and HeyA8-MDR cells. Columns, means of three experiments; bars, SE. *, P

    Article Snippet: The medium was exchanged after 48 hours with increasing concentrations of docetaxel (obtained from Aventis Pharma, Bridgewater, NJ) dissolved in ethanol (final concentration range, 0.1–10,000 nmol/L prepared in medium) or cisplatin (purchased from LKT Laboratories, Inc., St. Paul, MN) dissolved in water.

    Techniques: Activity Assay

    Bax protein expression induced by 0 nM and 20 nM docetaxel in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 (A) and H460 (B) cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, Western blot analyses were performed to detect the expression of Bax. The graph depicts mean±SEM for three independent determinations of optical density of the Bax Western blot bands. * Compared to control cells with incubation of 0 nM docetaxel, p

    Journal: Yonsei Medical Journal

    Article Title: Knockdown of the M2 Isoform of Pyruvate Kinase (PKM2) with shRNA Enhances the Effect of Docetaxel in Human NSCLC Cell Lines In Vitro

    doi: 10.3349/ymj.2016.57.6.1312

    Figure Lengend Snippet: Bax protein expression induced by 0 nM and 20 nM docetaxel in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 (A) and H460 (B) cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, Western blot analyses were performed to detect the expression of Bax. The graph depicts mean±SEM for three independent determinations of optical density of the Bax Western blot bands. * Compared to control cells with incubation of 0 nM docetaxel, p

    Article Snippet: Docetaxel, the reference drug, was purchased from Jiangsu Hengrui Medicine Co., Ltd.

    Techniques: Expressing, shRNA, Transfection, Incubation, Western Blot

    Effects of shRNA PKM2 and docetaxel on apoptosis. 48 hrs after transfection, all A549 and H460 cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with docetaxel at concentrations ranging from 0 to 25 nM for up to 48 hrs, and apoptosis was evaluated using Annexin V-FITC/PI staining and flow cytometry analysis. In both A549 and H460 cells, apoptosis was detected at two different time points after incubation, 24 hrs (A and B) and 48 hrs (C and D). Results are presented as mean±SEM of three separate experiments conducted in duplicate. * Compared to control cells, p

    Journal: Yonsei Medical Journal

    Article Title: Knockdown of the M2 Isoform of Pyruvate Kinase (PKM2) with shRNA Enhances the Effect of Docetaxel in Human NSCLC Cell Lines In Vitro

    doi: 10.3349/ymj.2016.57.6.1312

    Figure Lengend Snippet: Effects of shRNA PKM2 and docetaxel on apoptosis. 48 hrs after transfection, all A549 and H460 cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with docetaxel at concentrations ranging from 0 to 25 nM for up to 48 hrs, and apoptosis was evaluated using Annexin V-FITC/PI staining and flow cytometry analysis. In both A549 and H460 cells, apoptosis was detected at two different time points after incubation, 24 hrs (A and B) and 48 hrs (C and D). Results are presented as mean±SEM of three separate experiments conducted in duplicate. * Compared to control cells, p

    Article Snippet: Docetaxel, the reference drug, was purchased from Jiangsu Hengrui Medicine Co., Ltd.

    Techniques: shRNA, Transfection, Incubation, Staining, Flow Cytometry, Cytometry

    The effect of shRNA-PKM2 and docetaxel or the combined treatment of both on the viability of A549 and H460 cells. 48 hrs after transfection, all A549 and H460 cells, untransfected (Control), PKM2 shRNA-transfected (PKM2 shRNA) or control shRNA-transfected (Control shRNA), were all cultured with different concentrations of docetaxel, ranging from 0 to 25 nM, for up to 72 hrs. (A and B) Cells were incubated with docetaxel at 0 nm for 72 hrs. The cell viability was detected at 24 hrs, 48 hrs and 72 hrs after incubation. (C and D) Cells were incubated with docetaxel at concentrations ranging from 0 nm to 25 nm for 72 hrs. Cell viability was quantified using Cell Counting Kit-8 and expressed as the percentage of the viability of control cells (0 h). Results are presented as mean±SEM of three separate experiments conducted in duplicate. * Compared to control cells with the same dose of docetaxel incubation at the same time point, p

    Journal: Yonsei Medical Journal

    Article Title: Knockdown of the M2 Isoform of Pyruvate Kinase (PKM2) with shRNA Enhances the Effect of Docetaxel in Human NSCLC Cell Lines In Vitro

    doi: 10.3349/ymj.2016.57.6.1312

    Figure Lengend Snippet: The effect of shRNA-PKM2 and docetaxel or the combined treatment of both on the viability of A549 and H460 cells. 48 hrs after transfection, all A549 and H460 cells, untransfected (Control), PKM2 shRNA-transfected (PKM2 shRNA) or control shRNA-transfected (Control shRNA), were all cultured with different concentrations of docetaxel, ranging from 0 to 25 nM, for up to 72 hrs. (A and B) Cells were incubated with docetaxel at 0 nm for 72 hrs. The cell viability was detected at 24 hrs, 48 hrs and 72 hrs after incubation. (C and D) Cells were incubated with docetaxel at concentrations ranging from 0 nm to 25 nm for 72 hrs. Cell viability was quantified using Cell Counting Kit-8 and expressed as the percentage of the viability of control cells (0 h). Results are presented as mean±SEM of three separate experiments conducted in duplicate. * Compared to control cells with the same dose of docetaxel incubation at the same time point, p

    Article Snippet: Docetaxel, the reference drug, was purchased from Jiangsu Hengrui Medicine Co., Ltd.

    Techniques: shRNA, Transfection, Cell Culture, Incubation, Cell Counting

    Effects of PKM2 shRNA and docetaxel on cell cycle distribution. 48 hrs after transfection, all A549 and H460 cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with docetaxel at concentrations ranging from 0 to 25 nM for up to 48 hrs, and the cell cycle distribution was evaluated using PI staining and flow cytometry analysis. In both A549 and H460 cells, cell cycle distribution was measured at two different time points after incubation, 24 hrs (A and B) and 48 hrs (C and D). Results are presented as mean±SEM of three separate experiments conducted in duplicate. * Compared to control cells, p

    Journal: Yonsei Medical Journal

    Article Title: Knockdown of the M2 Isoform of Pyruvate Kinase (PKM2) with shRNA Enhances the Effect of Docetaxel in Human NSCLC Cell Lines In Vitro

    doi: 10.3349/ymj.2016.57.6.1312

    Figure Lengend Snippet: Effects of PKM2 shRNA and docetaxel on cell cycle distribution. 48 hrs after transfection, all A549 and H460 cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with docetaxel at concentrations ranging from 0 to 25 nM for up to 48 hrs, and the cell cycle distribution was evaluated using PI staining and flow cytometry analysis. In both A549 and H460 cells, cell cycle distribution was measured at two different time points after incubation, 24 hrs (A and B) and 48 hrs (C and D). Results are presented as mean±SEM of three separate experiments conducted in duplicate. * Compared to control cells, p

    Article Snippet: Docetaxel, the reference drug, was purchased from Jiangsu Hengrui Medicine Co., Ltd.

    Techniques: shRNA, Transfection, Incubation, Staining, Flow Cytometry, Cytometry

    Representative analysis of apoptosis induced by 20 nM docetaxel in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 and H460 cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, and apoptosis was evaluated using Annexin V-FITC/PI staining and flow cytometry analysis. (A) PKM2-shRNA transfected A549 cells treated with 20 nM DOC. (B) shRNA-control transfected A549 cells treated with 20 nM DOC. (C) Non-transfected A549 cells treated with 20 nM DOC. (D) PKM2-shRNA transfected H460 cells treated with 20 nM DOC. (E) shRNA-control transfected H460 cells treated with 20 nM DOC. (F) Non-transfected H460 cells treated with 20 nM DOC. Bottom right quadrant; cells stained mainly by Annexin V (early apoptotic cells); top right quadrant, cells stained by both PI and Annexin V (late apopototic cells); top left quadrant, cells stained mainly by PI (necrotic cells); bottom left quadrant, cells negative for both Annexin V and PI. PKM2, the M2 isoform of pyruvate kinase; PI, propidium iodide.

    Journal: Yonsei Medical Journal

    Article Title: Knockdown of the M2 Isoform of Pyruvate Kinase (PKM2) with shRNA Enhances the Effect of Docetaxel in Human NSCLC Cell Lines In Vitro

    doi: 10.3349/ymj.2016.57.6.1312

    Figure Lengend Snippet: Representative analysis of apoptosis induced by 20 nM docetaxel in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 and H460 cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, and apoptosis was evaluated using Annexin V-FITC/PI staining and flow cytometry analysis. (A) PKM2-shRNA transfected A549 cells treated with 20 nM DOC. (B) shRNA-control transfected A549 cells treated with 20 nM DOC. (C) Non-transfected A549 cells treated with 20 nM DOC. (D) PKM2-shRNA transfected H460 cells treated with 20 nM DOC. (E) shRNA-control transfected H460 cells treated with 20 nM DOC. (F) Non-transfected H460 cells treated with 20 nM DOC. Bottom right quadrant; cells stained mainly by Annexin V (early apoptotic cells); top right quadrant, cells stained by both PI and Annexin V (late apopototic cells); top left quadrant, cells stained mainly by PI (necrotic cells); bottom left quadrant, cells negative for both Annexin V and PI. PKM2, the M2 isoform of pyruvate kinase; PI, propidium iodide.

    Article Snippet: Docetaxel, the reference drug, was purchased from Jiangsu Hengrui Medicine Co., Ltd.

    Techniques: shRNA, Transfection, Incubation, Staining, Flow Cytometry, Cytometry

    p21 protein expression induced by 0 nM and 20 nM docetaxel in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 (A) and H460 (B) cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, Western blot analyses were performed to detect the expression of p21. The graph depicts mean±SEM for three independent determinations of optical density of the p21 Western blot bands. * Compared to control cells with incubation of 0 nM docetaxel, p

    Journal: Yonsei Medical Journal

    Article Title: Knockdown of the M2 Isoform of Pyruvate Kinase (PKM2) with shRNA Enhances the Effect of Docetaxel in Human NSCLC Cell Lines In Vitro

    doi: 10.3349/ymj.2016.57.6.1312

    Figure Lengend Snippet: p21 protein expression induced by 0 nM and 20 nM docetaxel in PKM2-shRNA transfected, shRNA-control transfected or non-transfected cells at 48 hrs. 48 hrs after transfection, all A549 (A) and H460 (B) cells, PKM2 shRNA-transfected (PKM2 shRNA), control-shRNA transfected (Control shRNA) or non-transfected (Control), were incubated with 20 nM docetaxel for up to 48 hrs, Western blot analyses were performed to detect the expression of p21. The graph depicts mean±SEM for three independent determinations of optical density of the p21 Western blot bands. * Compared to control cells with incubation of 0 nM docetaxel, p

    Article Snippet: Docetaxel, the reference drug, was purchased from Jiangsu Hengrui Medicine Co., Ltd.

    Techniques: Expressing, shRNA, Transfection, Incubation, Western Blot

    Decay-corrected TACs of [ 11 C]docetaxel (%ID/ml versus time p.i.) in plasma and whole blood

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: Decay-corrected TACs of [ 11 C]docetaxel (%ID/ml versus time p.i.) in plasma and whole blood

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques:

    PET/CT images of [ 11 C]docetaxel in a 71-year-old male patient with metastatic malignant pleural mesothelioma. Mediastinal metastasis ( arrow ) on CT scan ( a ) with [ 11 C]docetaxel uptake on PET scan ( b ) and on PET/CT fusion image ( c )

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: PET/CT images of [ 11 C]docetaxel in a 71-year-old male patient with metastatic malignant pleural mesothelioma. Mediastinal metastasis ( arrow ) on CT scan ( a ) with [ 11 C]docetaxel uptake on PET scan ( b ) and on PET/CT fusion image ( c )

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques: Positron Emission Tomography, Computed Tomography

    Decay-corrected TACs of [ 11 C]docetaxel for seven different tumours (SUV max versus time p.i.) including mediastinal metastasis of malignant pleural mesothelioma ( a ), metastasis of NSCLC ( b , d ), primary tumour of NSCLC ( c , e , f ) and lung metastasis of prostate cancer ( g )

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: Decay-corrected TACs of [ 11 C]docetaxel for seven different tumours (SUV max versus time p.i.) including mediastinal metastasis of malignant pleural mesothelioma ( a ), metastasis of NSCLC ( b , d ), primary tumour of NSCLC ( c , e , f ) and lung metastasis of prostate cancer ( g )

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques:

    For organs with activity clearly exceeding background (e.g. liver) ROIs were defined on the PET scan with the highest uptake. PET/CT fusion image ( a ) and PET image ( b ) demonstrate high [ 11 C]docetaxel uptake in liver. The liver ROI as determined on the PET image is presented in green ( b )

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: For organs with activity clearly exceeding background (e.g. liver) ROIs were defined on the PET scan with the highest uptake. PET/CT fusion image ( a ) and PET image ( b ) demonstrate high [ 11 C]docetaxel uptake in liver. The liver ROI as determined on the PET image is presented in green ( b )

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques: Activity Assay, Positron Emission Tomography

    For organs with lower tracer uptake (e.g. lung), ROIs were drawn on the LD-CT scan and subsequently copied onto the PET scans. PET/CT fusion image ( a ) and PET image demonstrate low [ 11 C]docetaxel uptake in lungs. The lung ROIs were drawn on the CT image ( b ) and projected onto the PET image ( c ). The lung ROIs as determined on the LD-CT scan are presented in green ( b , c )

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: For organs with lower tracer uptake (e.g. lung), ROIs were drawn on the LD-CT scan and subsequently copied onto the PET scans. PET/CT fusion image ( a ) and PET image demonstrate low [ 11 C]docetaxel uptake in lungs. The lung ROIs were drawn on the CT image ( b ) and projected onto the PET image ( c ). The lung ROIs as determined on the LD-CT scan are presented in green ( b , c )

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques: Computed Tomography, Positron Emission Tomography

    Decay-corrected TACs of [ 11 C]docetaxel (%ID versus time p.i.) ( n = 7) in a liver, b ULI and gall bladder, c brain, lung, heart contents and myocardial wall, and d urinary bladder, spleen, kidney and red marrow. Vertical bars indicate standard deviation

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: Decay-corrected TACs of [ 11 C]docetaxel (%ID versus time p.i.) ( n = 7) in a liver, b ULI and gall bladder, c brain, lung, heart contents and myocardial wall, and d urinary bladder, spleen, kidney and red marrow. Vertical bars indicate standard deviation

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques: Standard Deviation

    Maximum intensity projections of the biodistribution of [ 11 C]docetaxel in a 71-year-old male patient with metastatic malignant pleural mesothelioma. The four successive whole-body PET scans (0–6, 8–19, 23–39 and 42–63 min p.i.) demonstrate high liver uptake at all time points and low uptake in brain and normal lung. On the fourth scan, [ 11 C]docetaxel uptake is observed in intestine

    Journal: European Journal of Nuclear Medicine and Molecular Imaging

    Article Title: Biodistribution and radiation dosimetry of 11C-labelled docetaxel in cancer patients

    doi: 10.1007/s00259-010-1489-y

    Figure Lengend Snippet: Maximum intensity projections of the biodistribution of [ 11 C]docetaxel in a 71-year-old male patient with metastatic malignant pleural mesothelioma. The four successive whole-body PET scans (0–6, 8–19, 23–39 and 42–63 min p.i.) demonstrate high liver uptake at all time points and low uptake in brain and normal lung. On the fourth scan, [ 11 C]docetaxel uptake is observed in intestine

    Article Snippet: For the preparation of the precursor, the drug docetaxel was obtained from Green Plantchem Company Ltd. (Hangzhou, China).

    Techniques: Positron Emission Tomography

    Typical mass spectra obtained from a the spot of the docetaxel using ( a ) α-cyano-4-hydroxycinnamic acid (CHCA); ( b ) 2,5-dihydroxy benzoic acid (DHB); ( c ) 4-nitroaniline (4NA); and ( d ) 6-aza-2-thiothymine (ATT). M indicates docetaxel.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: Typical mass spectra obtained from a the spot of the docetaxel using ( a ) α-cyano-4-hydroxycinnamic acid (CHCA); ( b ) 2,5-dihydroxy benzoic acid (DHB); ( c ) 4-nitroaniline (4NA); and ( d ) 6-aza-2-thiothymine (ATT). M indicates docetaxel.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    Relationship between the signal intensities of [M + Na] + and [M + K] + . Black and gray bars mean that additive agent or no additive agent, respectively. M indicated docetaxel.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: Relationship between the signal intensities of [M + Na] + and [M + K] + . Black and gray bars mean that additive agent or no additive agent, respectively. M indicated docetaxel.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    Typical mass spectra obtained from docetaxel using ( a ) 4NA; ( b ) 4NA/NaY; ( c ) ATT; ( d ) ATT/NaY with no additive agents. M indicates docetaxel.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: Typical mass spectra obtained from docetaxel using ( a ) 4NA; ( b ) 4NA/NaY; ( c ) ATT; ( d ) ATT/NaY with no additive agents. M indicates docetaxel.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    Relationship between S/N ratio of the docetaxel peak using zeolite matrix with no additive agents, at the indicated concentrations and laser pulse energies.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: Relationship between S/N ratio of the docetaxel peak using zeolite matrix with no additive agents, at the indicated concentrations and laser pulse energies.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    The average of peak intensity of the sodium-adducted ion of docetaxel using zeolite matrix with 4NA or ATT with no additive agents.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: The average of peak intensity of the sodium-adducted ion of docetaxel using zeolite matrix with 4NA or ATT with no additive agents.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    Typical mass spectra obtained from the prostate cancer cells (PC-3) cells administered 100 μM docetaxel using ( a ) the zeolite matrix; ATT/NaY or ( b ) the conventional matrix, ATT.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: Typical mass spectra obtained from the prostate cancer cells (PC-3) cells administered 100 μM docetaxel using ( a ) the zeolite matrix; ATT/NaY or ( b ) the conventional matrix, ATT.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    Ion image of ( a ) m / z 829.5 [D + Na] + ; and ( b ) m / z 563.3 [P + H] + obtained from PC-3 cells administered 50 μM docetaxel and 10 μM protoporphyrin IX (PpIX) using the zeolite matrix, ATT/NaY with the droplet method.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: Ion image of ( a ) m / z 829.5 [D + Na] + ; and ( b ) m / z 563.3 [P + H] + obtained from PC-3 cells administered 50 μM docetaxel and 10 μM protoporphyrin IX (PpIX) using the zeolite matrix, ATT/NaY with the droplet method.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques:

    S/N ratio of the peak of [M + Na] + peak at 0, 10 and 100 μM docetaxel.

    Journal: International Journal of Molecular Sciences

    Article Title: Development of Laser Ionization Techniques for Evaluation of the Effect of Cancer Drugs Using Imaging Mass Spectrometry

    doi: 10.3390/ijms150711234

    Figure Lengend Snippet: S/N ratio of the peak of [M + Na] + peak at 0, 10 and 100 μM docetaxel.

    Article Snippet: Docetaxel trihydrate was purchased from Wako (Osaka, Japan).

    Techniques: