Structured Review

Taconic Biosciences hct116 p53
MiR-339-5p increases <t>p53</t> protein accumulation and its transcriptional activity in response to stress by negatively regulating MDM2 in human colorectal cancer cells (A) MiR-339-5p decreased MDM2 protein levels and increased the p53 protein accumulation and transcriptional activity toward p21 in response to stress in <t>HCT116</t> cells. (B) MiR-339-5p increased the p53 transcriptional activity toward p21, Puma and Fas in response to stress in HCT116 cells. In A and B: HCT116 p53+/+ and p53−/− cells were transfected with miR-339-5p mimic or miR-con. At 24 h after transfection, cells were treated with 5-FU (50 μM) for 8 h, and analyzed by western-blot (A) and Taqman real-time PCR (B), respectively. The mRNA levels of all genes were normalized to actin. The mRNA levels of genes in untreated cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). (C) MiR-339-5p decreased MDM2 protein levels and increased p53, p21 protein levels in response to stress in RKO cells. RKO p53+/+ and RKO p53−/− cells were treated with 5-FU and analyzed as described in A.
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1) Product Images from "MicroRNA-339-5p inhibits colorectal tumorigenesis through regulation of the MDM2/p53 signaling"

Article Title: MicroRNA-339-5p inhibits colorectal tumorigenesis through regulation of the MDM2/p53 signaling

Journal: Oncotarget

doi:

MiR-339-5p increases p53 protein accumulation and its transcriptional activity in response to stress by negatively regulating MDM2 in human colorectal cancer cells (A) MiR-339-5p decreased MDM2 protein levels and increased the p53 protein accumulation and transcriptional activity toward p21 in response to stress in HCT116 cells. (B) MiR-339-5p increased the p53 transcriptional activity toward p21, Puma and Fas in response to stress in HCT116 cells. In A and B: HCT116 p53+/+ and p53−/− cells were transfected with miR-339-5p mimic or miR-con. At 24 h after transfection, cells were treated with 5-FU (50 μM) for 8 h, and analyzed by western-blot (A) and Taqman real-time PCR (B), respectively. The mRNA levels of all genes were normalized to actin. The mRNA levels of genes in untreated cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). (C) MiR-339-5p decreased MDM2 protein levels and increased p53, p21 protein levels in response to stress in RKO cells. RKO p53+/+ and RKO p53−/− cells were treated with 5-FU and analyzed as described in A.
Figure Legend Snippet: MiR-339-5p increases p53 protein accumulation and its transcriptional activity in response to stress by negatively regulating MDM2 in human colorectal cancer cells (A) MiR-339-5p decreased MDM2 protein levels and increased the p53 protein accumulation and transcriptional activity toward p21 in response to stress in HCT116 cells. (B) MiR-339-5p increased the p53 transcriptional activity toward p21, Puma and Fas in response to stress in HCT116 cells. In A and B: HCT116 p53+/+ and p53−/− cells were transfected with miR-339-5p mimic or miR-con. At 24 h after transfection, cells were treated with 5-FU (50 μM) for 8 h, and analyzed by western-blot (A) and Taqman real-time PCR (B), respectively. The mRNA levels of all genes were normalized to actin. The mRNA levels of genes in untreated cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). (C) MiR-339-5p decreased MDM2 protein levels and increased p53, p21 protein levels in response to stress in RKO cells. RKO p53+/+ and RKO p53−/− cells were treated with 5-FU and analyzed as described in A.

Techniques Used: Activity Assay, Transfection, Western Blot, Real-time Polymerase Chain Reaction

MiR-339-5p inhibits the growth of HCT116 xenograft tumors in a largely p53-dependent manner (A, B) MiR-339-5p inhibited the growth of HCT116 xenograft tumors in nude mice in a largely p53-dependent manner. Xenograft tumors were established by s.c. injection of HCT116 p53+/+ and HCT116 p53−/− cells into nude mice. When tumor volumes reached ~60mm 3 , tumors were injected with miR-339-5p mimic or miR-con once every two days for 6 times. (A) Representative tumors were photographed at 12 days after the first treatment with miR-339-5p mimic or miR-con. Scale bar: 10 mm. (B) Upper panels: The growth curves of HCT116 p53+/+ and p53−/− tumors after miR-339-5p injection. The relative volumes of the tumors before treatment at day 0 were designated as 1. Lower panel: The fold reduction of tumor volumes by miR-339-5p injection in both HCT116 p53+/+ and p53−/− tumors. Data are presented as mean ± SD (n=12 for each group). *: p
Figure Legend Snippet: MiR-339-5p inhibits the growth of HCT116 xenograft tumors in a largely p53-dependent manner (A, B) MiR-339-5p inhibited the growth of HCT116 xenograft tumors in nude mice in a largely p53-dependent manner. Xenograft tumors were established by s.c. injection of HCT116 p53+/+ and HCT116 p53−/− cells into nude mice. When tumor volumes reached ~60mm 3 , tumors were injected with miR-339-5p mimic or miR-con once every two days for 6 times. (A) Representative tumors were photographed at 12 days after the first treatment with miR-339-5p mimic or miR-con. Scale bar: 10 mm. (B) Upper panels: The growth curves of HCT116 p53+/+ and p53−/− tumors after miR-339-5p injection. The relative volumes of the tumors before treatment at day 0 were designated as 1. Lower panel: The fold reduction of tumor volumes by miR-339-5p injection in both HCT116 p53+/+ and p53−/− tumors. Data are presented as mean ± SD (n=12 for each group). *: p

Techniques Used: Mouse Assay, Injection

MiR-339-5p negatively regulates MDM2 levels in human colorectal cells through binding to human MDM2 3′-UTR (A) MiR-339-5p decreased MDM2 protein levels and increased p53 protein levels in human colorectal cancer HCT116 and RKO cells. HCT116 p53+/+, HCT116 p53−/−, RKO p53+/+ and RKO p53−/− cells were transfected with miR-339-5p mimic or scrambled miRNA control (miR-con), and the MDM2 and p53 protein levels were measured at 24 h after transfection by western-blot assays. (B) MiR-339-5p decreased MDM2 mRNA levels in HCT116 and RKO cells. The MDM2 mRNA levels were measured by Taqman real-time PCR in cells transfected with miR-339-5p mimic or miR-con, and normalized with actin. The levels of the MDM2 mRNA in control cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). #: p
Figure Legend Snippet: MiR-339-5p negatively regulates MDM2 levels in human colorectal cells through binding to human MDM2 3′-UTR (A) MiR-339-5p decreased MDM2 protein levels and increased p53 protein levels in human colorectal cancer HCT116 and RKO cells. HCT116 p53+/+, HCT116 p53−/−, RKO p53+/+ and RKO p53−/− cells were transfected with miR-339-5p mimic or scrambled miRNA control (miR-con), and the MDM2 and p53 protein levels were measured at 24 h after transfection by western-blot assays. (B) MiR-339-5p decreased MDM2 mRNA levels in HCT116 and RKO cells. The MDM2 mRNA levels were measured by Taqman real-time PCR in cells transfected with miR-339-5p mimic or miR-con, and normalized with actin. The levels of the MDM2 mRNA in control cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). #: p

Techniques Used: Binding Assay, Transfection, Western Blot, Real-time Polymerase Chain Reaction

MiR-339-5p enhances p53-mediated apoptosis and senescence in response to stress (A) MiR-339-5p enhanced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 5-FU (300 μM), and apoptosis were measured by Annexin V staining in a flow cytometer at 24 or 36 h after treatment. (B) The miR-339-5p inhibitor reduced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with 5-FU and analyzed as described in A. (C) MiR-339-5p enhanced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 100 nM Doxorubicin for 3 days before cellular senescence was measured by SA-β-gal staining. (D) The miR-339-5p inhibitor reduced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with Doxorubicin and analyzed as described in C. In C and D: The upper panels are represented images of SA-β-gal staining. In A-D: Data are presented as mean ± SD (n = 3). #: p
Figure Legend Snippet: MiR-339-5p enhances p53-mediated apoptosis and senescence in response to stress (A) MiR-339-5p enhanced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 5-FU (300 μM), and apoptosis were measured by Annexin V staining in a flow cytometer at 24 or 36 h after treatment. (B) The miR-339-5p inhibitor reduced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with 5-FU and analyzed as described in A. (C) MiR-339-5p enhanced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 100 nM Doxorubicin for 3 days before cellular senescence was measured by SA-β-gal staining. (D) The miR-339-5p inhibitor reduced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with Doxorubicin and analyzed as described in C. In C and D: The upper panels are represented images of SA-β-gal staining. In A-D: Data are presented as mean ± SD (n = 3). #: p

Techniques Used: Transfection, Staining, Flow Cytometry, Cytometry

MiR-339-5p inhibits the migration and invasion of colorectal cancer cells in a largely p53-dependent manner (A) MiR-339-5p inhibited the migration of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into chambers for migration assays. Left panels: Representative images of migrated cells; Right panel: Quantifications of average number of migrated cells per field. (B) MiR-339-5p inhibited the invasion of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into matrigel-coated chambers for invasion assays. Left panels: Representative images of invading cells; Right panel: Quantifications of average number of invading cells per field. (C, D) The miR-339-5p inhibitor promoted the migration (C) and invasion (D) of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p inhibitor or miR-con inhibitor were used for migration and invasion assays as described in A and B, respectively. Data are presented as mean ± SD (n=3). #: p
Figure Legend Snippet: MiR-339-5p inhibits the migration and invasion of colorectal cancer cells in a largely p53-dependent manner (A) MiR-339-5p inhibited the migration of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into chambers for migration assays. Left panels: Representative images of migrated cells; Right panel: Quantifications of average number of migrated cells per field. (B) MiR-339-5p inhibited the invasion of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into matrigel-coated chambers for invasion assays. Left panels: Representative images of invading cells; Right panel: Quantifications of average number of invading cells per field. (C, D) The miR-339-5p inhibitor promoted the migration (C) and invasion (D) of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p inhibitor or miR-con inhibitor were used for migration and invasion assays as described in A and B, respectively. Data are presented as mean ± SD (n=3). #: p

Techniques Used: Migration, Transfection

2) Product Images from "Urokinase-Type Plasminogen Activator in Inflammatory Cell Recruitment and Host Defense against Pneumocystis carinii in Mice"

Article Title: Urokinase-Type Plasminogen Activator in Inflammatory Cell Recruitment and Host Defense against Pneumocystis carinii in Mice

Journal: Infection and Immunity

doi:

Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P
Figure Legend Snippet: Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P

Techniques Used: Knock-Out, Mouse Assay

Histology of P. carinii infection. Lungs of wild-type mice inoculated with P. carinii demonstrated perivascular and peribronchiolar mononuclear cell infiltrates (arrow) (A) but no P. carinii organisms (C). Lungs of uPA knockout mice inoculated with P. carinii demonstrated markedly reduced mononuclear cell inflammation but contained alveolar exudate indicative of P. carinii infection (arrows) (B), confirmed by the presence of P. carinii organisms (arrowheads) (D). (A and B) H E stain; magnification, ×200. (C and D) Gomori methenamine silver stain; magnification, ×400.
Figure Legend Snippet: Histology of P. carinii infection. Lungs of wild-type mice inoculated with P. carinii demonstrated perivascular and peribronchiolar mononuclear cell infiltrates (arrow) (A) but no P. carinii organisms (C). Lungs of uPA knockout mice inoculated with P. carinii demonstrated markedly reduced mononuclear cell inflammation but contained alveolar exudate indicative of P. carinii infection (arrows) (B), confirmed by the presence of P. carinii organisms (arrowheads) (D). (A and B) H E stain; magnification, ×200. (C and D) Gomori methenamine silver stain; magnification, ×400.

Techniques Used: Infection, Mouse Assay, Knock-Out, Staining, Silver Staining

Intensity of P. carinii infection. Wild-type ( n = 9) and uPA knockout ( n = 10) mice were inoculated intratracheally with P. carinii organisms, and the intensity of infection was scored 4 weeks after inoculation, using a histologic grading scale. Wild-type mice cleared the P. carinii inoculum, with only a single mouse demonstrating several individual, widely scattered P. carinii cysts. Conversely, uPA knockout mice were uniformly heavily infected with P. carinii , with all mice demonstrating grade 3 or grade 4 infections. Data represent medians obtained from two individual experiments. ∗, P
Figure Legend Snippet: Intensity of P. carinii infection. Wild-type ( n = 9) and uPA knockout ( n = 10) mice were inoculated intratracheally with P. carinii organisms, and the intensity of infection was scored 4 weeks after inoculation, using a histologic grading scale. Wild-type mice cleared the P. carinii inoculum, with only a single mouse demonstrating several individual, widely scattered P. carinii cysts. Conversely, uPA knockout mice were uniformly heavily infected with P. carinii , with all mice demonstrating grade 3 or grade 4 infections. Data represent medians obtained from two individual experiments. ∗, P

Techniques Used: Infection, Knock-Out, Mouse Assay

Cellular constituents of BAL fluid. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were characterized 4 weeks after inoculation. Total numbers of lymphocytes (A), macrophages (B), and neutrophils (C) are indicated. Data represent means ± SEM; ∗, P
Figure Legend Snippet: Cellular constituents of BAL fluid. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were characterized 4 weeks after inoculation. Total numbers of lymphocytes (A), macrophages (B), and neutrophils (C) are indicated. Data represent means ± SEM; ∗, P

Techniques Used: Knock-Out, Mouse Assay

3) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m
Figure Legend Snippet: Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m

Techniques Used: In Vitro, Multiple Displacement Amplification

4) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

5) Product Images from "DENND2B activates Rab13 at the leading edge of migrating cells and promotes metastatic behavior"

Article Title: DENND2B activates Rab13 at the leading edge of migrating cells and promotes metastatic behavior

Journal: The Journal of Cell Biology

doi: 10.1083/jcb.201407068

Rab13 knockdown reduces tumor growth and migration in vivo. (A) 1833TR cells stably expressing luciferase and either a control vector or shRNAs targeting Rab13 were injected into the tail veins of mice. The panel shows representative bioluminescence images of mice immediately after injection (day 0) and 63 d after injection. See Fig. S3 for all mice used in the study. (B) Tumor take rates of injected mice at day 63. (C) Total bioluminescence of luciferase-positive mice was quantified and normalized to day 0 (mean ± SEM from one independent experiment in which n = 9 for control mice and shRNA 2 and n = 8 for shRNA 1; two-way ANOVA with Tukey’s post-test; *, P
Figure Legend Snippet: Rab13 knockdown reduces tumor growth and migration in vivo. (A) 1833TR cells stably expressing luciferase and either a control vector or shRNAs targeting Rab13 were injected into the tail veins of mice. The panel shows representative bioluminescence images of mice immediately after injection (day 0) and 63 d after injection. See Fig. S3 for all mice used in the study. (B) Tumor take rates of injected mice at day 63. (C) Total bioluminescence of luciferase-positive mice was quantified and normalized to day 0 (mean ± SEM from one independent experiment in which n = 9 for control mice and shRNA 2 and n = 8 for shRNA 1; two-way ANOVA with Tukey’s post-test; *, P

Techniques Used: Migration, In Vivo, Stable Transfection, Expressing, Luciferase, Plasmid Preparation, Injection, Mouse Assay, shRNA

6) Product Images from "Phospho-Ibuprofen (MDC-917) Is a Novel Agent against Colon Cancer: Efficacy, Metabolism, and Pharmacokinetics in Mouse Models"

Article Title: Phospho-Ibuprofen (MDC-917) Is a Novel Agent against Colon Cancer: Efficacy, Metabolism, and Pharmacokinetics in Mouse Models

Journal: The Journal of Pharmacology and Experimental Therapeutics

doi: 10.1124/jpet.111.180224

Effect of phospho-ibuprofen on colon cancer cell growth in vitro and in vivo. Top, IC 50 values for colon cancer cells treated with PI or ibuprofen for 24 h. Bottom, PI inhibits the growth of SW480 human colon cancer xenografts in nude mice. The study
Figure Legend Snippet: Effect of phospho-ibuprofen on colon cancer cell growth in vitro and in vivo. Top, IC 50 values for colon cancer cells treated with PI or ibuprofen for 24 h. Bottom, PI inhibits the growth of SW480 human colon cancer xenografts in nude mice. The study

Techniques Used: In Vitro, In Vivo, Mouse Assay

Correlation of PI metabolite levels with tumor volume. Mice with SW480 colon cancer xenografts were sacrificed 1 h after the last dosing, and ibuprofen levels in mouse blood and tumors were determined as described under Materials and Methods . A significant
Figure Legend Snippet: Correlation of PI metabolite levels with tumor volume. Mice with SW480 colon cancer xenografts were sacrificed 1 h after the last dosing, and ibuprofen levels in mouse blood and tumors were determined as described under Materials and Methods . A significant

Techniques Used: Mouse Assay

7) Product Images from "KIBRA (WWC1) Is a Metastasis Suppressor Gene Affected by Chromosome 5q Loss in Triple-Negative Breast Cancer"

Article Title: KIBRA (WWC1) Is a Metastasis Suppressor Gene Affected by Chromosome 5q Loss in Triple-Negative Breast Cancer

Journal: Cell Reports

doi: 10.1016/j.celrep.2018.02.095

Anti-tumorigenic Effects of KIBRA Are Associated with a Reduction in TAZ Protein Levels and Inhibition of TAZ Nuclear Localization (Ai–Aiii) Subcellular localization of YAP/TAZ in MDA-MB-231 cells expressing wild-type KIBRA (KIBRA-WT), ΔWW1/2-KIBRA, or control (pLVX-GFP) in response to increasing matrix tension (0.3 to 17 kPa). Scale bars, 20 μm. (Aiv) Quantification of nuclear to cytoplasmic YAP/TAZ ratios under conditions of soft (0.3 kPa) or stiff (17 kPa) matrix or a collagen-coated glass coverslip (70 GPa) (n = 3, mean ± SD). (Bi and Bii) Representative images and quantification of YAP/TAZ localization in A1005 cells in response to matrix tension as in (A). (C) Western blot showing YAP phosphorylation and TAZ protein levels in MDA-MB-231 and A1005 cells with or without KIBRA. (Di) Western blots confirming transfection of constitutively active YAP or TAZ in A1005 and MDA-MB-231 cells with KIBRA. The empty pCMV vector is a negative control. The arrows indicate tagged (top) and endogenous (bottom) proteins. (Dii) Quantification of SFE relative to empty vector for cells in (Di) (n = 3, ± SEM). (Diii) Representative tumorsphere images. Scale bars, 400 μm. (Ei) Immunohistochemistry (IHC) showing YAP and TAZ subcellular localization in A1005 orthotopic tumors with or without KIBRA . Scale bars, 100 μm. (Eii and Eiii) Percentage of cells with positive nuclear staining (ii) and mean optical density (OD) of nuclear staining (iii) for YAP and TAZ in ten fields of view, 6 to 10 sections per condition (mean ± SEM). .
Figure Legend Snippet: Anti-tumorigenic Effects of KIBRA Are Associated with a Reduction in TAZ Protein Levels and Inhibition of TAZ Nuclear Localization (Ai–Aiii) Subcellular localization of YAP/TAZ in MDA-MB-231 cells expressing wild-type KIBRA (KIBRA-WT), ΔWW1/2-KIBRA, or control (pLVX-GFP) in response to increasing matrix tension (0.3 to 17 kPa). Scale bars, 20 μm. (Aiv) Quantification of nuclear to cytoplasmic YAP/TAZ ratios under conditions of soft (0.3 kPa) or stiff (17 kPa) matrix or a collagen-coated glass coverslip (70 GPa) (n = 3, mean ± SD). (Bi and Bii) Representative images and quantification of YAP/TAZ localization in A1005 cells in response to matrix tension as in (A). (C) Western blot showing YAP phosphorylation and TAZ protein levels in MDA-MB-231 and A1005 cells with or without KIBRA. (Di) Western blots confirming transfection of constitutively active YAP or TAZ in A1005 and MDA-MB-231 cells with KIBRA. The empty pCMV vector is a negative control. The arrows indicate tagged (top) and endogenous (bottom) proteins. (Dii) Quantification of SFE relative to empty vector for cells in (Di) (n = 3, ± SEM). (Diii) Representative tumorsphere images. Scale bars, 400 μm. (Ei) Immunohistochemistry (IHC) showing YAP and TAZ subcellular localization in A1005 orthotopic tumors with or without KIBRA . Scale bars, 100 μm. (Eii and Eiii) Percentage of cells with positive nuclear staining (ii) and mean optical density (OD) of nuclear staining (iii) for YAP and TAZ in ten fields of view, 6 to 10 sections per condition (mean ± SEM). .

Techniques Used: Inhibition, Multiple Displacement Amplification, Expressing, Western Blot, Transfection, Plasmid Preparation, Negative Control, Immunohistochemistry, Staining

8) Product Images from "Obesity-linked phosphorylation of PPAR? by cdk5 is a direct target of the anti-diabetic PPAR? ligands"

Article Title: Obesity-linked phosphorylation of PPAR? by cdk5 is a direct target of the anti-diabetic PPAR? ligands

Journal: Nature

doi: 10.1038/nature09291

Specific fat cell gene dysregulation by the cdk5-mediated S273 phosphorylation of PPARγ a , In vitro CDK assays performed using cdk5/p35 with either wild type (WT) or S273A mutated PPARγ. b , Phosphorylation of PPARγ in differentiated 3T3-L1 adipocytes stimulated with TNF-α for the indicated times. c , Phosphorylation of PPARγ in cells expressing scrambled or CDK5 shRNA stimulated with indicated cytokines. NT, no treatment. d , Staining of PPARγ-null fibroblasts expressing WT or S273A mutant PPARγ with Oil-Red-O. e , Gene expression in these cells was analyzed by real-time quantitative PCR (qPCR) for expression of various genes (n=3). f , mRNA expression in transplanted fat pads was analyzed by qPCR (n=5) (Error bars are s.e.m.; * p
Figure Legend Snippet: Specific fat cell gene dysregulation by the cdk5-mediated S273 phosphorylation of PPARγ a , In vitro CDK assays performed using cdk5/p35 with either wild type (WT) or S273A mutated PPARγ. b , Phosphorylation of PPARγ in differentiated 3T3-L1 adipocytes stimulated with TNF-α for the indicated times. c , Phosphorylation of PPARγ in cells expressing scrambled or CDK5 shRNA stimulated with indicated cytokines. NT, no treatment. d , Staining of PPARγ-null fibroblasts expressing WT or S273A mutant PPARγ with Oil-Red-O. e , Gene expression in these cells was analyzed by real-time quantitative PCR (qPCR) for expression of various genes (n=3). f , mRNA expression in transplanted fat pads was analyzed by qPCR (n=5) (Error bars are s.e.m.; * p

Techniques Used: In Vitro, Expressing, shRNA, Staining, Mutagenesis, Real-time Polymerase Chain Reaction

Anti-diabetic PPARγ ligands block CDK5-mediated phosphorylation of PPARγ a , TNF-α-induced phosphorylation of PPARγ in 3T3-L1 adipocytes expressing either WT or Q286P mutant of PPARγ treated with rosiglitazone and/or GW9662. b and c , In vitro CDK5 assay with either rosiglitazone or MRL24. d , Transcriptional activity of a PPAR-derived reporter gene in response to rosiglitazone or MRL24 (n=3). e , Microarray analysis of differentiated PPARγ-null fibroblasts expressing WT (NT, rosiglitazone or MRL24 treated) or S273A mutant PPARγ. f , mRNA expression of genes regulated by the phosphorylation of PPARγ in epididymal fat tissue of mice on either chow or HFD for 13 weeks (n=5). Error bars are s.e.m.; * p
Figure Legend Snippet: Anti-diabetic PPARγ ligands block CDK5-mediated phosphorylation of PPARγ a , TNF-α-induced phosphorylation of PPARγ in 3T3-L1 adipocytes expressing either WT or Q286P mutant of PPARγ treated with rosiglitazone and/or GW9662. b and c , In vitro CDK5 assay with either rosiglitazone or MRL24. d , Transcriptional activity of a PPAR-derived reporter gene in response to rosiglitazone or MRL24 (n=3). e , Microarray analysis of differentiated PPARγ-null fibroblasts expressing WT (NT, rosiglitazone or MRL24 treated) or S273A mutant PPARγ. f , mRNA expression of genes regulated by the phosphorylation of PPARγ in epididymal fat tissue of mice on either chow or HFD for 13 weeks (n=5). Error bars are s.e.m.; * p

Techniques Used: Blocking Assay, Expressing, Mutagenesis, In Vitro, Activity Assay, Derivative Assay, Microarray, Mouse Assay

9) Product Images from "NK Cells Promote Th-17 Mediated Corneal Barrier Disruption in Dry Eye"

Article Title: NK Cells Promote Th-17 Mediated Corneal Barrier Disruption in Dry Eye

Journal: PLoS ONE

doi: 10.1371/journal.pone.0036822

Location of intra-epithelial lymphocytes in conjunctiva before and after desiccating stress. Representative digital pictures of immunohistochemical staining of CD4 + , CD8α + , CD103 + , γδTCR + , NK + in the conjunctivae of nonstressed (NS) C57BL/6 mice and after desiccating stress for 5 or 10 days (DS5, DS10). Inset in γδTCR row shows γδTCR in skin, which was used as positive control. Original magnification 40X; scale bar 20 µm. Number insets represent cell counts in the goblet cell rich of the conjunctiva in immunostained tissue sections in conjunctival epithelium of C57BL/6 mice. Data represents mean ± SD of cells/mm. Experiments were repeated three times with two mice per group per experiment. * indicates p
Figure Legend Snippet: Location of intra-epithelial lymphocytes in conjunctiva before and after desiccating stress. Representative digital pictures of immunohistochemical staining of CD4 + , CD8α + , CD103 + , γδTCR + , NK + in the conjunctivae of nonstressed (NS) C57BL/6 mice and after desiccating stress for 5 or 10 days (DS5, DS10). Inset in γδTCR row shows γδTCR in skin, which was used as positive control. Original magnification 40X; scale bar 20 µm. Number insets represent cell counts in the goblet cell rich of the conjunctiva in immunostained tissue sections in conjunctival epithelium of C57BL/6 mice. Data represents mean ± SD of cells/mm. Experiments were repeated three times with two mice per group per experiment. * indicates p

Techniques Used: Immunohistochemistry, Staining, Mouse Assay, Positive Control

10) Product Images from "A Notch1 Ectodomain Construct Inhibits Endothelial Notch Signaling, Tumor Growth, and Angiogenesis"

Article Title: A Notch1 Ectodomain Construct Inhibits Endothelial Notch Signaling, Tumor Growth, and Angiogenesis

Journal: Cancer research

doi: 10.1158/0008-5472.CAN-07-6499

Notch1 decoy expression disrupts angiogenesis and impairs tumor viability in human NGP xenografts. We have previously reported that human neuroblastoma xenografts in mice have a mature, hierarchical vasculature that is relatively resistant to VEGF blockade
Figure Legend Snippet: Notch1 decoy expression disrupts angiogenesis and impairs tumor viability in human NGP xenografts. We have previously reported that human neuroblastoma xenografts in mice have a mature, hierarchical vasculature that is relatively resistant to VEGF blockade

Techniques Used: Expressing, Mouse Assay

11) Product Images from "Integrative discovery of treatments for high-risk neuroblastoma"

Article Title: Integrative discovery of treatments for high-risk neuroblastoma

Journal: Nature Communications

doi: 10.1038/s41467-019-13817-8

GW405833 reduces neuroblastoma growth in vivo. a Mice were engrafted with \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$15 \ \times 1{0}^{6}$$\end{document} 15 × 1 0 6 SK-N-BE(2) cells s.c. and randomized to receive a daily i.p. injection of GW (45 mg/kg; n = 10) or vehicle ( n = 12) for 8 days, starting at the appearance of palpable tumours of \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${ > }0.2\,{\mathrm{cm}}^{3}$$\end{document} > 0.2 cm 3 . b GW405833 significantly impaired the growth of established human tumours (hierarchical linear model). c Point comparison of day 8 tumour volume. d Tumour volume increase from day 0 to day 8. e Post-mortem tumour weight after 8 days of treatment. f Cell proliferation marker MKI67, counted using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 4). g Apoptosis marker cleaved PARP, counted in using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 3). h Representative images of tumour histology (HE), MKI67 and c-PARP localization, bar = 100 μm. Statistics: b Mean, 95% confidence interval, p -value computed from a mixed effects model and corrected for multiple testing using bonferroni correction. c – g Mean, standard deviation, Student’s t -test.
Figure Legend Snippet: GW405833 reduces neuroblastoma growth in vivo. a Mice were engrafted with \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$15 \ \times 1{0}^{6}$$\end{document} 15 × 1 0 6 SK-N-BE(2) cells s.c. and randomized to receive a daily i.p. injection of GW (45 mg/kg; n = 10) or vehicle ( n = 12) for 8 days, starting at the appearance of palpable tumours of \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${ > }0.2\,{\mathrm{cm}}^{3}$$\end{document} > 0.2 cm 3 . b GW405833 significantly impaired the growth of established human tumours (hierarchical linear model). c Point comparison of day 8 tumour volume. d Tumour volume increase from day 0 to day 8. e Post-mortem tumour weight after 8 days of treatment. f Cell proliferation marker MKI67, counted using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 4). g Apoptosis marker cleaved PARP, counted in using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 3). h Representative images of tumour histology (HE), MKI67 and c-PARP localization, bar = 100 μm. Statistics: b Mean, 95% confidence interval, p -value computed from a mixed effects model and corrected for multiple testing using bonferroni correction. c – g Mean, standard deviation, Student’s t -test.

Techniques Used: In Vivo, Mouse Assay, Injection, Marker, Standard Deviation

12) Product Images from "Characterization of two mouse models of metastatic pheochromocytoma using bioluminescence imaging"

Article Title: Characterization of two mouse models of metastatic pheochromocytoma using bioluminescence imaging

Journal: Cancer Letters

doi: 10.1016/j.canlet.2011.10.019

Longitudinal subcutaneous tumor monitoring. MTT-luc (1 × 10 6 cells) cells were injected subcutaneously in the left flank of Nude mice (n= 6). (A). Representative images from the same mouse and taken weekly after implantation, are presented. (B)
Figure Legend Snippet: Longitudinal subcutaneous tumor monitoring. MTT-luc (1 × 10 6 cells) cells were injected subcutaneously in the left flank of Nude mice (n= 6). (A). Representative images from the same mouse and taken weekly after implantation, are presented. (B)

Techniques Used: MTT Assay, Injection, Mouse Assay

Histopathological analysis in metastatic sites. Representative histological sections of liver and lung metastasis from MTT-luc cells. The presence of metastatic tumor cells initially detected in vivo and ex vivo by BLI imaging was confirmed by histopathological
Figure Legend Snippet: Histopathological analysis in metastatic sites. Representative histological sections of liver and lung metastasis from MTT-luc cells. The presence of metastatic tumor cells initially detected in vivo and ex vivo by BLI imaging was confirmed by histopathological

Techniques Used: MTT Assay, In Vivo, Ex Vivo, Imaging

(A) Bioluminescence signal of the murine pheochromocytoma MTT-luc cells in vitro . MTT-luc cells were diluted from 4,000 to 8 cells, plated in duplicate in a 96 well plate and imaged for 20 seconds after addition of luciferin to the media. Media alone
Figure Legend Snippet: (A) Bioluminescence signal of the murine pheochromocytoma MTT-luc cells in vitro . MTT-luc cells were diluted from 4,000 to 8 cells, plated in duplicate in a 96 well plate and imaged for 20 seconds after addition of luciferin to the media. Media alone

Techniques Used: MTT Assay, In Vitro

3.2 Detection and non-invasive BLI monitoring of MTT-luc cells in an experimental metastasis model
Figure Legend Snippet: 3.2 Detection and non-invasive BLI monitoring of MTT-luc cells in an experimental metastasis model

Techniques Used: MTT Assay

13) Product Images from "Validation of the Hsp70-Bag3 Protein-Protein Interaction as a Potential Therapeutic Target in Cancer"

Article Title: Validation of the Hsp70-Bag3 Protein-Protein Interaction as a Potential Therapeutic Target in Cancer

Journal: Molecular cancer therapeutics

doi: 10.1158/1535-7163.MCT-14-0650

JG-98 is active in MCF7 and HeLa xenograft models. JG-98 treatment of mice with either (A) MCF7 cells or (B) HeLa cells xenografted. See the materials and methods for experimental details. Arrows indicate treatments with JG-98. (C) Treated samples from
Figure Legend Snippet: JG-98 is active in MCF7 and HeLa xenograft models. JG-98 treatment of mice with either (A) MCF7 cells or (B) HeLa cells xenografted. See the materials and methods for experimental details. Arrows indicate treatments with JG-98. (C) Treated samples from

Techniques Used: Mouse Assay

14) Product Images from "TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis"

Article Title: TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis

Journal: Oncogene

doi: 10.1038/s41388-017-0043-9

DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P
Figure Legend Snippet: DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P

Techniques Used: In Vivo, Stable Transfection, Expressing, shRNA, Sequencing, Mouse Assay, Injection

DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.
Figure Legend Snippet: DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.

Techniques Used: Transduction, Cell Culture, Incubation, Recombinant, SDS Page, Staining

15) Product Images from "KIBRA (WWC1) Is a Metastasis Suppressor Gene Affected by Chromosome 5q Loss in Triple-Negative Breast Cancer"

Article Title: KIBRA (WWC1) Is a Metastasis Suppressor Gene Affected by Chromosome 5q Loss in Triple-Negative Breast Cancer

Journal: Cell Reports

doi: 10.1016/j.celrep.2018.02.095

Anti-tumorigenic Effects of KIBRA Are Associated with a Reduction in TAZ Protein Levels and Inhibition of TAZ Nuclear Localization (Ai–Aiii) Subcellular localization of YAP/TAZ in MDA-MB-231 cells expressing wild-type KIBRA (KIBRA-WT), ΔWW1/2-KIBRA, or control (pLVX-GFP) in response to increasing matrix tension (0.3 to 17 kPa). Scale bars, 20 μm. (Aiv) Quantification of nuclear to cytoplasmic YAP/TAZ ratios under conditions of soft (0.3 kPa) or stiff (17 kPa) matrix or a collagen-coated glass coverslip (70 GPa) (n = 3, mean ± SD). (Bi and Bii) Representative images and quantification of YAP/TAZ localization in A1005 cells in response to matrix tension as in (A). (C) Western blot showing YAP phosphorylation and TAZ protein levels in MDA-MB-231 and A1005 cells with or without KIBRA. (Di) Western blots confirming transfection of constitutively active YAP or TAZ in A1005 and MDA-MB-231 cells with KIBRA. The empty pCMV vector is a negative control. The arrows indicate tagged (top) and endogenous (bottom) proteins. (Dii) Quantification of SFE relative to empty vector for cells in (Di) (n = 3, ± SEM). (Diii) Representative tumorsphere images. Scale bars, 400 μm. (Ei) Immunohistochemistry (IHC) showing YAP and TAZ subcellular localization in A1005 orthotopic tumors with or without KIBRA . Scale bars, 100 μm. (Eii and Eiii) Percentage of cells with positive nuclear staining (ii) and mean optical density (OD) of nuclear staining (iii) for YAP and TAZ in ten fields of view, 6 to 10 sections per condition (mean ± SEM). See also Figure S5 .
Figure Legend Snippet: Anti-tumorigenic Effects of KIBRA Are Associated with a Reduction in TAZ Protein Levels and Inhibition of TAZ Nuclear Localization (Ai–Aiii) Subcellular localization of YAP/TAZ in MDA-MB-231 cells expressing wild-type KIBRA (KIBRA-WT), ΔWW1/2-KIBRA, or control (pLVX-GFP) in response to increasing matrix tension (0.3 to 17 kPa). Scale bars, 20 μm. (Aiv) Quantification of nuclear to cytoplasmic YAP/TAZ ratios under conditions of soft (0.3 kPa) or stiff (17 kPa) matrix or a collagen-coated glass coverslip (70 GPa) (n = 3, mean ± SD). (Bi and Bii) Representative images and quantification of YAP/TAZ localization in A1005 cells in response to matrix tension as in (A). (C) Western blot showing YAP phosphorylation and TAZ protein levels in MDA-MB-231 and A1005 cells with or without KIBRA. (Di) Western blots confirming transfection of constitutively active YAP or TAZ in A1005 and MDA-MB-231 cells with KIBRA. The empty pCMV vector is a negative control. The arrows indicate tagged (top) and endogenous (bottom) proteins. (Dii) Quantification of SFE relative to empty vector for cells in (Di) (n = 3, ± SEM). (Diii) Representative tumorsphere images. Scale bars, 400 μm. (Ei) Immunohistochemistry (IHC) showing YAP and TAZ subcellular localization in A1005 orthotopic tumors with or without KIBRA . Scale bars, 100 μm. (Eii and Eiii) Percentage of cells with positive nuclear staining (ii) and mean optical density (OD) of nuclear staining (iii) for YAP and TAZ in ten fields of view, 6 to 10 sections per condition (mean ± SEM). See also Figure S5 .

Techniques Used: Inhibition, Multiple Displacement Amplification, Expressing, Western Blot, Transfection, Plasmid Preparation, Negative Control, Immunohistochemistry, Staining

16) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m
Figure Legend Snippet: Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m

Techniques Used: In Vitro, Multiple Displacement Amplification

17) Product Images from "A Notch1 Ectodomain Construct Inhibits Endothelial Notch Signaling, Tumor Growth, and Angiogenesis"

Article Title: A Notch1 Ectodomain Construct Inhibits Endothelial Notch Signaling, Tumor Growth, and Angiogenesis

Journal: Cancer research

doi: 10.1158/0008-5472.CAN-07-6499

Notch1 decoy inhibits angiogenesis and s.c. tumor growth of Mm5MT-FGF4 tumors in mice. A, Mm5MT-FGF4 tumor growth in soft agar is little affected by expression of Fc or N1 decoy. B, tumor volumes of Mm5MT-FGF4-X and Mm5MT-FGF4-Fc differ significantly
Figure Legend Snippet: Notch1 decoy inhibits angiogenesis and s.c. tumor growth of Mm5MT-FGF4 tumors in mice. A, Mm5MT-FGF4 tumor growth in soft agar is little affected by expression of Fc or N1 decoy. B, tumor volumes of Mm5MT-FGF4-X and Mm5MT-FGF4-Fc differ significantly

Techniques Used: Mouse Assay, Expressing

Notch1 decoy or compound E blocks Notch4-mediated HUVEC extensions. A, ectopic expression of Notch4 induces morphogenetic changes by HUVECs cultured on fibrin gel. HUVECs were transduced with Ad-Notch4 at 30 m.o.i. and Ad-GFP at 10 m.o.i. to mark-infected
Figure Legend Snippet: Notch1 decoy or compound E blocks Notch4-mediated HUVEC extensions. A, ectopic expression of Notch4 induces morphogenetic changes by HUVECs cultured on fibrin gel. HUVECs were transduced with Ad-Notch4 at 30 m.o.i. and Ad-GFP at 10 m.o.i. to mark-infected

Techniques Used: Expressing, Cell Culture, Transduction, Infection

Role of Notch signaling in VEGF-dependent in vivo angiogenesis. Inhibition of KP1/VEGF-induced angiogenesis with Notch1 decoy in mouse DAS assay. Representative photographs. Top, subcutaneous VEGF-induced angiogenesis with control COL1 ( left ) and KP1/VEGF
Figure Legend Snippet: Role of Notch signaling in VEGF-dependent in vivo angiogenesis. Inhibition of KP1/VEGF-induced angiogenesis with Notch1 decoy in mouse DAS assay. Representative photographs. Top, subcutaneous VEGF-induced angiogenesis with control COL1 ( left ) and KP1/VEGF

Techniques Used: In Vivo, Inhibition

Notch1 decoy expression disrupts angiogenesis and impairs tumor viability in human NGP xenografts. We have previously reported that human neuroblastoma xenografts in mice have a mature, hierarchical vasculature that is relatively resistant to VEGF blockade
Figure Legend Snippet: Notch1 decoy expression disrupts angiogenesis and impairs tumor viability in human NGP xenografts. We have previously reported that human neuroblastoma xenografts in mice have a mature, hierarchical vasculature that is relatively resistant to VEGF blockade

Techniques Used: Expressing, Mouse Assay

Notch1 decoy inhibits activation of Notch signaling stimulated by Notch ligands. A, schematic of Notch1 decoy containing the 36 endothelial growth factor repeats of rat Notch1 fused to human Fc. B, Western blotting to detect secreted Notch1 decoy in conditioned
Figure Legend Snippet: Notch1 decoy inhibits activation of Notch signaling stimulated by Notch ligands. A, schematic of Notch1 decoy containing the 36 endothelial growth factor repeats of rat Notch1 fused to human Fc. B, Western blotting to detect secreted Notch1 decoy in conditioned

Techniques Used: Activation Assay, Western Blot

18) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m
Figure Legend Snippet: Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m

Techniques Used: In Vitro, Multiple Displacement Amplification

19) Product Images from "Novel multi-targeted ErbB family inhibitor afatinib blocks EGF-induced signaling and induces apoptosis in neuroblastoma"

Article Title: Novel multi-targeted ErbB family inhibitor afatinib blocks EGF-induced signaling and induces apoptosis in neuroblastoma

Journal: Oncotarget

doi: 10.18632/oncotarget.13657

Afatinib blocks EGF-induced phosphorylation of EGFR, AKT and S6 in NB cells A-B. Six NB cell lines (IMR-32, NGP, NB-19, SK-N-AS, SH-SY5Y, and LA-N-6) were starved for 16 hrs in serum-free medium before exposed to afatinib (10 μM) treatment for 1 hr. Then the cells were stimulated with or without 100 ng/ml hEGF for 10 min. Cells were then collected and subjected to SDS-PAGE, immunoblotted with the indicated antibodies, respectively. β-Actin was used as a loading control in all samples.
Figure Legend Snippet: Afatinib blocks EGF-induced phosphorylation of EGFR, AKT and S6 in NB cells A-B. Six NB cell lines (IMR-32, NGP, NB-19, SK-N-AS, SH-SY5Y, and LA-N-6) were starved for 16 hrs in serum-free medium before exposed to afatinib (10 μM) treatment for 1 hr. Then the cells were stimulated with or without 100 ng/ml hEGF for 10 min. Cells were then collected and subjected to SDS-PAGE, immunoblotted with the indicated antibodies, respectively. β-Actin was used as a loading control in all samples.

Techniques Used: SDS Page

Afatinib induces apoptosis by blocking PI3K/AKT/mTOR signaling in an orthotopic xenograft NB mouse model The mice bearing SH-SY5Y-luciferase cells xenografted tumors for four weeks were treated with either afatinib (25 mg/kg) or an equal volume of DMSO by i.p. injection daily for three days. Four hours after the last treatment, the mice were sacrificed and the tumors were harvested and lysed for immunoblotting with the indicated antibodies. β-Actin was used as a loading control.
Figure Legend Snippet: Afatinib induces apoptosis by blocking PI3K/AKT/mTOR signaling in an orthotopic xenograft NB mouse model The mice bearing SH-SY5Y-luciferase cells xenografted tumors for four weeks were treated with either afatinib (25 mg/kg) or an equal volume of DMSO by i.p. injection daily for three days. Four hours after the last treatment, the mice were sacrificed and the tumors were harvested and lysed for immunoblotting with the indicated antibodies. β-Actin was used as a loading control.

Techniques Used: Blocking Assay, Mouse Assay, Luciferase, Injection

Afatinib shows cytotoxic effect on NB cells A. Six NB cell lines (IMR-32, NGP, NB-19, SK-N-AS, SH-SY5Y and LA-N-6) were treated with increasing concentrations of afatinib for 72 hrs. Cell viability was then assessed by a CCK-8 assay. Data were presented as mean ± SD. P
Figure Legend Snippet: Afatinib shows cytotoxic effect on NB cells A. Six NB cell lines (IMR-32, NGP, NB-19, SK-N-AS, SH-SY5Y and LA-N-6) were treated with increasing concentrations of afatinib for 72 hrs. Cell viability was then assessed by a CCK-8 assay. Data were presented as mean ± SD. P

Techniques Used: CCK-8 Assay

Afatinib suppresses the anchorage-independent growth of NB cells A. A panel of six NB cell lines were seeded in six-well plates with indicated concentrations of afatinib in soft agar, and grown for 2 to 3 weeks, followed by staining with crystal violet for 4 hrs and the photos were taken. B. Colonies were counted and colony numbers were presented as mean ± SD. P
Figure Legend Snippet: Afatinib suppresses the anchorage-independent growth of NB cells A. A panel of six NB cell lines were seeded in six-well plates with indicated concentrations of afatinib in soft agar, and grown for 2 to 3 weeks, followed by staining with crystal violet for 4 hrs and the photos were taken. B. Colonies were counted and colony numbers were presented as mean ± SD. P

Techniques Used: Staining

Afatinib induces apoptosis in NB cells A-F. IMR-32, NGP, NB-19, SK-N-AS and SH-SY5Y and LA-N-6 cells were treated with afatinib (10 μM or 20 μM) for various time points (0-16 hrs). At the end of treatment, cells were harvested and cell lysates were subjected to SDS-PAGE, and then immunoblotted with the indicated antibodies. β-Actin was used as a loading control.
Figure Legend Snippet: Afatinib induces apoptosis in NB cells A-F. IMR-32, NGP, NB-19, SK-N-AS and SH-SY5Y and LA-N-6 cells were treated with afatinib (10 μM or 20 μM) for various time points (0-16 hrs). At the end of treatment, cells were harvested and cell lysates were subjected to SDS-PAGE, and then immunoblotted with the indicated antibodies. β-Actin was used as a loading control.

Techniques Used: SDS Page

Afatinib enhances doxorubicin-induced cytotoxicity in NB cells A. Six cell lines were seeded in 96-well plates and were incubated with doxorubicin at the indicated concentrations with or without afatinib (2 μM) for 48 hrs. Cell viability was then measured by CCK-8 assay. Data were represented as mean ± SD. P
Figure Legend Snippet: Afatinib enhances doxorubicin-induced cytotoxicity in NB cells A. Six cell lines were seeded in 96-well plates and were incubated with doxorubicin at the indicated concentrations with or without afatinib (2 μM) for 48 hrs. Cell viability was then measured by CCK-8 assay. Data were represented as mean ± SD. P

Techniques Used: Incubation, CCK-8 Assay

20) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m
Figure Legend Snippet: Plasma stability and in vitro cytotoxicity. a Stability in human plasma. b Stability in mouse plasma. Cell killing potency in the breast cancer cell lines KPL-4 ( c ), JIMT-1 ( d ), and MDA-MB-231 ( e ). We tested unconjugated N297A anti-HER2 mAb (black cross), VCit ADC ( 3a , blue circle), SVCit ADC ( 3b , orange triangle), EVCit ADC ( 3c , green square), non-cleavable ADC ( 4 , red inversed triangle), and isotype control ADC containing EVCit ( 5 , magenta diamond, non-targeting control). All assays were performed in quadruplicate. Error bars represent s.e.m

Techniques Used: In Vitro, Multiple Displacement Amplification

21) Product Images from "Photoacoustic imaging to localize indeterminate pulmonary nodules: A preclinical study"

Article Title: Photoacoustic imaging to localize indeterminate pulmonary nodules: A preclinical study

Journal: PLoS ONE

doi: 10.1371/journal.pone.0231488

US and PA images and corresponding PA spectrogram from injected ICG-agar into healthy subcutaneous tissue at 0hr (A), 24hr (B), and 48hr (C) Fig 4D US and PA image and spectrogram obtained from ICG agar injected deep to subcutaneous tissue H460 tumor (48hr).
Figure Legend Snippet: US and PA images and corresponding PA spectrogram from injected ICG-agar into healthy subcutaneous tissue at 0hr (A), 24hr (B), and 48hr (C) Fig 4D US and PA image and spectrogram obtained from ICG agar injected deep to subcutaneous tissue H460 tumor (48hr).

Techniques Used: Injection

The images of H460 cells without ICG (A-C) and with ICG incubation (D-F) using confocal microscopy.
Figure Legend Snippet: The images of H460 cells without ICG (A-C) and with ICG incubation (D-F) using confocal microscopy.

Techniques Used: Incubation, Confocal Microscopy

22) Product Images from "CD4 T Cells Are the Only Lymphocytes Needed To Protect Mice against Rotavirus Shedding after Intranasal Immunization with a Chimeric VP6 Protein and the Adjuvant LT(R192G)"

Article Title: CD4 T Cells Are the Only Lymphocytes Needed To Protect Mice against Rotavirus Shedding after Intranasal Immunization with a Chimeric VP6 Protein and the Adjuvant LT(R192G)

Journal: Journal of Virology

doi: 10.1128/JVI.76.2.560-568.2002

Rotavirus shedding in Rag-2 mice after adoptive transfer of purified CD4 T cells from naive mice. A group of five naive BALB/c mice were sacrificed, and their splenic CD4 T cells were column purified and sorted twice. Specified quantities of these cells were then injected (i.p.) into groups of four Rag-2 mice that were chronically shedding high levels of rotavirus after oral inoculation of 1,000 SD 50 of wild-type EDIM 26 days earlier (day 0). Monitoring for rotavirus antigen shedding was continued until day 61.
Figure Legend Snippet: Rotavirus shedding in Rag-2 mice after adoptive transfer of purified CD4 T cells from naive mice. A group of five naive BALB/c mice were sacrificed, and their splenic CD4 T cells were column purified and sorted twice. Specified quantities of these cells were then injected (i.p.) into groups of four Rag-2 mice that were chronically shedding high levels of rotavirus after oral inoculation of 1,000 SD 50 of wild-type EDIM 26 days earlier (day 0). Monitoring for rotavirus antigen shedding was continued until day 61.

Techniques Used: Mouse Assay, Adoptive Transfer Assay, Purification, Injection

Rotavirus shedding in Rag-2 mice after adoptive transfer of purified CD4 T cells from VP6-immunized mice. A group of four BALB/c mice were immunized i.n. with two doses of MBP::VP6 and LT(R192G) and 4 weeks after the second dose were sacrificed. Their splenic CD4 T cells were column purified and sorted twice prior to i.p. injection of different quantities into groups of eight Rag-2 mice that were chronically shedding large quantities of rotavirus after oral inoculation of 1,000 SD 50 of wild-type EDIM 26 days earlier (day 0). EDIM shedding continued to be monitored daily in all groups until day 60. Starting on day 61, the two groups that received the larger numbers of CD4 T cells were depleted with anti-CD4 MAb injections for 4 consecutive days, and shedding of rotavirus antigen continued to be monitored until day 80.
Figure Legend Snippet: Rotavirus shedding in Rag-2 mice after adoptive transfer of purified CD4 T cells from VP6-immunized mice. A group of four BALB/c mice were immunized i.n. with two doses of MBP::VP6 and LT(R192G) and 4 weeks after the second dose were sacrificed. Their splenic CD4 T cells were column purified and sorted twice prior to i.p. injection of different quantities into groups of eight Rag-2 mice that were chronically shedding large quantities of rotavirus after oral inoculation of 1,000 SD 50 of wild-type EDIM 26 days earlier (day 0). EDIM shedding continued to be monitored daily in all groups until day 60. Starting on day 61, the two groups that received the larger numbers of CD4 T cells were depleted with anti-CD4 MAb injections for 4 consecutive days, and shedding of rotavirus antigen continued to be monitored until day 80.

Techniques Used: Mouse Assay, Adoptive Transfer Assay, Purification, Injection

FACS analysis of either B cells or CD8 or CD4 T cells obtained from the spleens, IELs, or lamina propria of Rag-2 mice 7 weeks after adoptive transfer of purified splenic CD4 T cells from VP6-immunized mice. Splenic CD4 T cells were obtained 4 weeks after the second i.n. immunization of BALB/c mice with MBP::VP6 and LT(R192G), column purified and sorted twice. These cells (2 × 10 5 ).
Figure Legend Snippet: FACS analysis of either B cells or CD8 or CD4 T cells obtained from the spleens, IELs, or lamina propria of Rag-2 mice 7 weeks after adoptive transfer of purified splenic CD4 T cells from VP6-immunized mice. Splenic CD4 T cells were obtained 4 weeks after the second i.n. immunization of BALB/c mice with MBP::VP6 and LT(R192G), column purified and sorted twice. These cells (2 × 10 5 ).

Techniques Used: FACS, Mouse Assay, Adoptive Transfer Assay, Purification

23) Product Images from "Vaccination with Recombinant N-Terminal Domain of Als1p Improves Survival during Murine Disseminated Candidiasis by Enhancing Cell-Mediated, Not Humoral, Immunity "

Article Title: Vaccination with Recombinant N-Terminal Domain of Als1p Improves Survival during Murine Disseminated Candidiasis by Enhancing Cell-Mediated, Not Humoral, Immunity

Journal: Infection and Immunity

doi: 10.1128/IAI.73.2.999-1005.2005

The rAls1p-N vaccine requires T cells, but not B cells, to induce protective immunity. Survival of B-cell-deficient, T-cell-deficient (nude), and congenic wild-type BALB/c control mice (seven or eight per group) was simultaneously assessed after vaccination with rAls1p-N plus adjuvant or with adjuvant alone. *, P
Figure Legend Snippet: The rAls1p-N vaccine requires T cells, but not B cells, to induce protective immunity. Survival of B-cell-deficient, T-cell-deficient (nude), and congenic wild-type BALB/c control mice (seven or eight per group) was simultaneously assessed after vaccination with rAls1p-N plus adjuvant or with adjuvant alone. *, P

Techniques Used: Mouse Assay

24) Product Images from "Discovery of 8-Cyclopentyl-2-[4-(4-methyl-piperazin-1-yl)-phenylamino]-7-oxo-7,8-dihydro-pyrido[2,3-d]pyrimidine-6-carbonitrile ( 7x) as a Potent Inhibitor of Cyclin-Dependent Kinase 4 (CDK4) and AMPK-Related Kinase 5 (ARK5)"

Article Title: Discovery of 8-Cyclopentyl-2-[4-(4-methyl-piperazin-1-yl)-phenylamino]-7-oxo-7,8-dihydro-pyrido[2,3-d]pyrimidine-6-carbonitrile ( 7x) as a Potent Inhibitor of Cyclin-Dependent Kinase 4 (CDK4) and AMPK-Related Kinase 5 (ARK5)

Journal: Journal of Medicinal Chemistry

doi: 10.1021/jm401073p

7x treatment induces apoptosis of breast carcinoma cell lines: MCF-7 (A) and MDA-MB-231 (B) human breast carcinoma cell lines were treated with increasing concentrations of 7x or PD-0332991 (control) for 24 h. Total cell lysates were subjected to Western blot analysis using a PARP-specific antibody. The cleaved protein, which is expressed in 7x treated cells, is indicative of apoptosis.
Figure Legend Snippet: 7x treatment induces apoptosis of breast carcinoma cell lines: MCF-7 (A) and MDA-MB-231 (B) human breast carcinoma cell lines were treated with increasing concentrations of 7x or PD-0332991 (control) for 24 h. Total cell lysates were subjected to Western blot analysis using a PARP-specific antibody. The cleaved protein, which is expressed in 7x treated cells, is indicative of apoptosis.

Techniques Used: Multiple Displacement Amplification, Western Blot

In vivo efficacy of 7x against subcutaneous breast tumor xenografts: MDA-MB-231 cells were orthotopically implanted into the mammary fat pad of 6–8 week old female nude mice ( n = 11 per group). Treatment was started when the average tumor volume reached 100 mm 3 . 7x (lactate salt dissolved in PBS) or vehicle was administered intraperitoneally every other day (Q2D). Tumor volumes (A) and body weights (B) were recorded every 2 days. All values represent mean ± SEM.
Figure Legend Snippet: In vivo efficacy of 7x against subcutaneous breast tumor xenografts: MDA-MB-231 cells were orthotopically implanted into the mammary fat pad of 6–8 week old female nude mice ( n = 11 per group). Treatment was started when the average tumor volume reached 100 mm 3 . 7x (lactate salt dissolved in PBS) or vehicle was administered intraperitoneally every other day (Q2D). Tumor volumes (A) and body weights (B) were recorded every 2 days. All values represent mean ± SEM.

Techniques Used: In Vivo, Multiple Displacement Amplification, Mouse Assay

7x treatment inhibits AKT phosphorylation: MDA-MB-231 human breast carcinoma cells were treated with increasing concentrations of 7x or PD-0332991 or LY294002 (2-morpholino-8-phenyl-4 H -chromen-4-one) (PI3Kinase inhibitor) for 24 h. Total cell lysates were subjected to Western blot analysis using antibodies directed against phosphorylated (Ser 473 ) and nonphosphorylated forms of AKT. GAPDH was used as a loading control.
Figure Legend Snippet: 7x treatment inhibits AKT phosphorylation: MDA-MB-231 human breast carcinoma cells were treated with increasing concentrations of 7x or PD-0332991 or LY294002 (2-morpholino-8-phenyl-4 H -chromen-4-one) (PI3Kinase inhibitor) for 24 h. Total cell lysates were subjected to Western blot analysis using antibodies directed against phosphorylated (Ser 473 ) and nonphosphorylated forms of AKT. GAPDH was used as a loading control.

Techniques Used: Multiple Displacement Amplification, Western Blot

Effect of 7x on cell cycle progression: MCF-7 (A) and MDA-MB-231 (B) human breast carcinoma cell lines were treated with increasing concentrations of 7x or PD-0332991 (control) for 24 h. The cells were then harvested, fixed, and stained with propidium iodide prior to flow cytometric analysis. The percentage of cells at each phase of the cell cycle was calculated and represented as % cells in each phase in the bar graph.
Figure Legend Snippet: Effect of 7x on cell cycle progression: MCF-7 (A) and MDA-MB-231 (B) human breast carcinoma cell lines were treated with increasing concentrations of 7x or PD-0332991 (control) for 24 h. The cells were then harvested, fixed, and stained with propidium iodide prior to flow cytometric analysis. The percentage of cells at each phase of the cell cycle was calculated and represented as % cells in each phase in the bar graph.

Techniques Used: Multiple Displacement Amplification, Staining, Flow Cytometry

(A) Inhibition of CDK4/cyclin D1 activity by 7x : 10 ng of recombinant CDK4/cyclin D1 complex was incubated with the indicated concentrations of 7x , flavopiridol (FP), or PD-0332991 for 30 min at room temperature. Kinase reactions were initiated by the addition of the substrate mixture (5 μM ATP, 10 μci γ 32 P -ATP, 10 mM MgCl 2 , and 1 μg recombinant RB substrate) and incubated at 30 °C for 20 min. The reactions were terminated by the addition of 2× Laemmli sample buffer and heated at 95 °C for 3 min. Proteins were resolved by 12% SDS-PAGE and the resulting gel subjected to autoradiography. (B) 7x inhibits RB phosphorylation at serine 780: An estrogen-dependent breast cancer cell line, MCF-7, and the triple negative (C) human breast carcinoma cell line, MDA-MB-231, were treated with increasing concentrations of 7x or PD-0332991 (control) for 24 h. Western blot analysis was performed using antibodies directed against phosphorylated (Ser 780 ) and nonphosphorylated forms of the retinoblastoma protein. Both 7x and PD-0332991 inhibit RB phosphorylation at Ser 780 , a known substrate of CDK4.
Figure Legend Snippet: (A) Inhibition of CDK4/cyclin D1 activity by 7x : 10 ng of recombinant CDK4/cyclin D1 complex was incubated with the indicated concentrations of 7x , flavopiridol (FP), or PD-0332991 for 30 min at room temperature. Kinase reactions were initiated by the addition of the substrate mixture (5 μM ATP, 10 μci γ 32 P -ATP, 10 mM MgCl 2 , and 1 μg recombinant RB substrate) and incubated at 30 °C for 20 min. The reactions were terminated by the addition of 2× Laemmli sample buffer and heated at 95 °C for 3 min. Proteins were resolved by 12% SDS-PAGE and the resulting gel subjected to autoradiography. (B) 7x inhibits RB phosphorylation at serine 780: An estrogen-dependent breast cancer cell line, MCF-7, and the triple negative (C) human breast carcinoma cell line, MDA-MB-231, were treated with increasing concentrations of 7x or PD-0332991 (control) for 24 h. Western blot analysis was performed using antibodies directed against phosphorylated (Ser 780 ) and nonphosphorylated forms of the retinoblastoma protein. Both 7x and PD-0332991 inhibit RB phosphorylation at Ser 780 , a known substrate of CDK4.

Techniques Used: Inhibition, Activity Assay, Recombinant, Incubation, SDS Page, Autoradiography, Multiple Displacement Amplification, Western Blot

25) Product Images from "Activation of IRF1 in Human Adipocytes Leads to Phenotypes Associated with Metabolic Disease"

Article Title: Activation of IRF1 in Human Adipocytes Leads to Phenotypes Associated with Metabolic Disease

Journal: Stem Cell Reports

doi: 10.1016/j.stemcr.2017.03.014

IRF1 Leads to Increased Inflammation In Vivo (A) 3T3-F442A cells were transduced with either rtTA alone, or human IRF1 (hIRF1) containing lentiviral particles, and were injected into 6-week-old nude mice (n = 4). Six weeks after injection, ectopic subcutaneous fat pads (dashed circles) were excised. (B) Ectopic fat explants (EXP) were stained with DAPI (blue) and probed with α-F4/80 (green) and α-PLIN1 (red). (C) F4/80-positive cells were counted relative to DAPI staining foci in hIRF1 explants versus rtTA explants and autologous WAT (see Figure S3 C). A minimum of 10 4 DAPI foci were counted for each experiment. (D) qPCR was used to assess expression of hIRF1 and inflammation-associated mouse genes in both explants and autologous WAT (n = 3). Error bars represent SD, experiments were performed in biological triplicates, and statistically significant p values are denoted by asterisks ( ∗ p ≤ 0.05, ∗∗ p ≤ 0.005).
Figure Legend Snippet: IRF1 Leads to Increased Inflammation In Vivo (A) 3T3-F442A cells were transduced with either rtTA alone, or human IRF1 (hIRF1) containing lentiviral particles, and were injected into 6-week-old nude mice (n = 4). Six weeks after injection, ectopic subcutaneous fat pads (dashed circles) were excised. (B) Ectopic fat explants (EXP) were stained with DAPI (blue) and probed with α-F4/80 (green) and α-PLIN1 (red). (C) F4/80-positive cells were counted relative to DAPI staining foci in hIRF1 explants versus rtTA explants and autologous WAT (see Figure S3 C). A minimum of 10 4 DAPI foci were counted for each experiment. (D) qPCR was used to assess expression of hIRF1 and inflammation-associated mouse genes in both explants and autologous WAT (n = 3). Error bars represent SD, experiments were performed in biological triplicates, and statistically significant p values are denoted by asterisks ( ∗ p ≤ 0.05, ∗∗ p ≤ 0.005).

Techniques Used: In Vivo, Transduction, Injection, Mouse Assay, Staining, Real-time Polymerase Chain Reaction, Expressing

26) Product Images from "Obesity-linked phosphorylation of PPAR? by cdk5 is a direct target of the anti-diabetic PPAR? ligands"

Article Title: Obesity-linked phosphorylation of PPAR? by cdk5 is a direct target of the anti-diabetic PPAR? ligands

Journal: Nature

doi: 10.1038/nature09291

CDK5-mediated phosphorylation of PPARγ is increased in fat tissues of high fat diet fed mice (HFD) a , White adipose tissue (epididymal) from mice on HFD for the indicated time was analyzed with phospho-S273 PPARγ and PPARγ antibodies. b , Epididymal (Epi.) or inguinal (Ing.) fat tissue from 13 weeks HFD mice was analyzed with phospho-S273 antibody.
Figure Legend Snippet: CDK5-mediated phosphorylation of PPARγ is increased in fat tissues of high fat diet fed mice (HFD) a , White adipose tissue (epididymal) from mice on HFD for the indicated time was analyzed with phospho-S273 PPARγ and PPARγ antibodies. b , Epididymal (Epi.) or inguinal (Ing.) fat tissue from 13 weeks HFD mice was analyzed with phospho-S273 antibody.

Techniques Used: Mouse Assay

Correlation between the inhibition of phosphorylation and improvement of insulin sensitivity by anti-diabetic PPARγ ligands a , Glucose-tolerance tests in 16-week HFD mice treated with vehicle, rosiglitazone or MRL24 (n=10). b , phosphorylation of PPARγ in WAT. c , The expression of gene sets regulated by PPARγ phosphorylation in WAT (Error bars are s.e.m.; * p
Figure Legend Snippet: Correlation between the inhibition of phosphorylation and improvement of insulin sensitivity by anti-diabetic PPARγ ligands a , Glucose-tolerance tests in 16-week HFD mice treated with vehicle, rosiglitazone or MRL24 (n=10). b , phosphorylation of PPARγ in WAT. c , The expression of gene sets regulated by PPARγ phosphorylation in WAT (Error bars are s.e.m.; * p

Techniques Used: Inhibition, Mouse Assay, Expressing

Specific fat cell gene dysregulation by the cdk5-mediated S273 phosphorylation of PPARγ a , In vitro CDK assays performed using cdk5/p35 with either wild type (WT) or S273A mutated PPARγ. b , Phosphorylation of PPARγ in differentiated 3T3-L1 adipocytes stimulated with TNF-α for the indicated times. c , Phosphorylation of PPARγ in cells expressing scrambled or CDK5 shRNA stimulated with indicated cytokines. NT, no treatment. d , Staining of PPARγ-null fibroblasts expressing WT or S273A mutant PPARγ with Oil-Red-O. e , Gene expression in these cells was analyzed by real-time quantitative PCR (qPCR) for expression of various genes (n=3). f , mRNA expression in transplanted fat pads was analyzed by qPCR (n=5) (Error bars are s.e.m.; * p
Figure Legend Snippet: Specific fat cell gene dysregulation by the cdk5-mediated S273 phosphorylation of PPARγ a , In vitro CDK assays performed using cdk5/p35 with either wild type (WT) or S273A mutated PPARγ. b , Phosphorylation of PPARγ in differentiated 3T3-L1 adipocytes stimulated with TNF-α for the indicated times. c , Phosphorylation of PPARγ in cells expressing scrambled or CDK5 shRNA stimulated with indicated cytokines. NT, no treatment. d , Staining of PPARγ-null fibroblasts expressing WT or S273A mutant PPARγ with Oil-Red-O. e , Gene expression in these cells was analyzed by real-time quantitative PCR (qPCR) for expression of various genes (n=3). f , mRNA expression in transplanted fat pads was analyzed by qPCR (n=5) (Error bars are s.e.m.; * p

Techniques Used: In Vitro, Expressing, shRNA, Staining, Mutagenesis, Real-time Polymerase Chain Reaction

Anti-diabetic PPARγ ligands block CDK5-mediated phosphorylation of PPARγ a , TNF-α-induced phosphorylation of PPARγ in 3T3-L1 adipocytes expressing either WT or Q286P mutant of PPARγ treated with rosiglitazone and/or GW9662. b and c , In vitro CDK5 assay with either rosiglitazone or MRL24. d , Transcriptional activity of a PPAR-derived reporter gene in response to rosiglitazone or MRL24 (n=3). e , Microarray analysis of differentiated PPARγ-null fibroblasts expressing WT (NT, rosiglitazone or MRL24 treated) or S273A mutant PPARγ. f , mRNA expression of genes regulated by the phosphorylation of PPARγ in epididymal fat tissue of mice on either chow or HFD for 13 weeks (n=5). Error bars are s.e.m.; * p
Figure Legend Snippet: Anti-diabetic PPARγ ligands block CDK5-mediated phosphorylation of PPARγ a , TNF-α-induced phosphorylation of PPARγ in 3T3-L1 adipocytes expressing either WT or Q286P mutant of PPARγ treated with rosiglitazone and/or GW9662. b and c , In vitro CDK5 assay with either rosiglitazone or MRL24. d , Transcriptional activity of a PPAR-derived reporter gene in response to rosiglitazone or MRL24 (n=3). e , Microarray analysis of differentiated PPARγ-null fibroblasts expressing WT (NT, rosiglitazone or MRL24 treated) or S273A mutant PPARγ. f , mRNA expression of genes regulated by the phosphorylation of PPARγ in epididymal fat tissue of mice on either chow or HFD for 13 weeks (n=5). Error bars are s.e.m.; * p

Techniques Used: Blocking Assay, Expressing, Mutagenesis, In Vitro, Activity Assay, Derivative Assay, Microarray, Mouse Assay

Differential HDX MS data for PPARγ-LBD ± rosiglitazone and MRL24 a , Histograms showing the percent reduction in HDX for Helix 3 (IRIFQGCQF), the β-sheet region (ISEGQGFMTRE), Helix 12 (QEIYKDLY) and the Helix 2-2’ link region containing the site of CDK5 phosphorylation (KTTDKSPFVIYDM). Values are calculated relative to the measured %D value for apo PPARγ-LBD (n=4; error bars are s.e.m.; ** p
Figure Legend Snippet: Differential HDX MS data for PPARγ-LBD ± rosiglitazone and MRL24 a , Histograms showing the percent reduction in HDX for Helix 3 (IRIFQGCQF), the β-sheet region (ISEGQGFMTRE), Helix 12 (QEIYKDLY) and the Helix 2-2’ link region containing the site of CDK5 phosphorylation (KTTDKSPFVIYDM). Values are calculated relative to the measured %D value for apo PPARγ-LBD (n=4; error bars are s.e.m.; ** p

Techniques Used: Mass Spectrometry

27) Product Images from "Pleiotrophin Gene Therapy for Peripheral Ischemia: Evaluation of Full-Length and Truncated Gene Variants"

Article Title: Pleiotrophin Gene Therapy for Peripheral Ischemia: Evaluation of Full-Length and Truncated Gene Variants

Journal: PLoS ONE

doi: 10.1371/journal.pone.0061413

No evidence of tumorigenesis from full-length PTN expression in myoblasts. A) When PTN myoblasts and lacZ myoblasts were plated at equal densities and grown past confluence, the cells proliferated and declined at equal rates regardless of PTN expression. B) There was no difference between the cell types in percent viability after contact inhibition. C) The positive control MDA-MB-231 breast cancer cell line induced subcutaneous tumors at 3/6 implantation sites after 4 weeks (black arrows), whereas implantation of PTN myoblasts resulted in no detectable tumor formation (0/6; white arrows) in this experiment, and 0/5 after 10 weeks in a separate experiment (not shown). 1×10 5 cells were injected in 10 µl PBS for anterior injections and 10 µl Matrigel for posterior injections. Animals are shown 4 weeks post injection. Tumor #1 = 1×0.8×0.5 cm; tumor #2 = 0.3×0.3×0.2 cm; tumor #3 = 0.7×0.9×0.5 cm.
Figure Legend Snippet: No evidence of tumorigenesis from full-length PTN expression in myoblasts. A) When PTN myoblasts and lacZ myoblasts were plated at equal densities and grown past confluence, the cells proliferated and declined at equal rates regardless of PTN expression. B) There was no difference between the cell types in percent viability after contact inhibition. C) The positive control MDA-MB-231 breast cancer cell line induced subcutaneous tumors at 3/6 implantation sites after 4 weeks (black arrows), whereas implantation of PTN myoblasts resulted in no detectable tumor formation (0/6; white arrows) in this experiment, and 0/5 after 10 weeks in a separate experiment (not shown). 1×10 5 cells were injected in 10 µl PBS for anterior injections and 10 µl Matrigel for posterior injections. Animals are shown 4 weeks post injection. Tumor #1 = 1×0.8×0.5 cm; tumor #2 = 0.3×0.3×0.2 cm; tumor #3 = 0.7×0.9×0.5 cm.

Techniques Used: Expressing, Inhibition, Positive Control, Multiple Displacement Amplification, Injection

28) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

29) Product Images from "Discovery of 8-Cyclopentyl-2-[4-(4-methyl-piperazin-1-yl)-phenylamino]-7-oxo-7,8-dihydro-pyrido[2,3-d]pyrimidine-6-carbonitrile ( 7x) as a Potent Inhibitor of Cyclin-Dependent Kinase 4 (CDK4) and AMPK-Related Kinase 5 (ARK5)"

Article Title: Discovery of 8-Cyclopentyl-2-[4-(4-methyl-piperazin-1-yl)-phenylamino]-7-oxo-7,8-dihydro-pyrido[2,3-d]pyrimidine-6-carbonitrile ( 7x) as a Potent Inhibitor of Cyclin-Dependent Kinase 4 (CDK4) and AMPK-Related Kinase 5 (ARK5)

Journal: Journal of Medicinal Chemistry

doi: 10.1021/jm401073p

In vivo efficacy of 7x against subcutaneous breast tumor xenografts: MDA-MB-231 cells were orthotopically implanted into the mammary fat pad of 6–8 week old female nude mice ( n = 11 per group). Treatment was started when the average tumor volume reached 100 mm 3 . 7x (lactate salt dissolved in PBS) or vehicle was administered intraperitoneally every other day (Q2D). Tumor volumes (A) and body weights (B) were recorded every 2 days. All values represent mean ± SEM.
Figure Legend Snippet: In vivo efficacy of 7x against subcutaneous breast tumor xenografts: MDA-MB-231 cells were orthotopically implanted into the mammary fat pad of 6–8 week old female nude mice ( n = 11 per group). Treatment was started when the average tumor volume reached 100 mm 3 . 7x (lactate salt dissolved in PBS) or vehicle was administered intraperitoneally every other day (Q2D). Tumor volumes (A) and body weights (B) were recorded every 2 days. All values represent mean ± SEM.

Techniques Used: In Vivo, Multiple Displacement Amplification, Mouse Assay

30) Product Images from "Obesity-linked phosphorylation of PPAR? by cdk5 is a direct target of the anti-diabetic PPAR? ligands"

Article Title: Obesity-linked phosphorylation of PPAR? by cdk5 is a direct target of the anti-diabetic PPAR? ligands

Journal: Nature

doi: 10.1038/nature09291

CDK5-mediated phosphorylation of PPARγ is increased in fat tissues of high fat diet fed mice (HFD) a , White adipose tissue (epididymal) from mice on HFD for the indicated time was analyzed with phospho-S273 PPARγ and PPARγ antibodies. b , Epididymal (Epi.) or inguinal (Ing.) fat tissue from 13 weeks HFD mice was analyzed with phospho-S273 antibody.
Figure Legend Snippet: CDK5-mediated phosphorylation of PPARγ is increased in fat tissues of high fat diet fed mice (HFD) a , White adipose tissue (epididymal) from mice on HFD for the indicated time was analyzed with phospho-S273 PPARγ and PPARγ antibodies. b , Epididymal (Epi.) or inguinal (Ing.) fat tissue from 13 weeks HFD mice was analyzed with phospho-S273 antibody.

Techniques Used: Mouse Assay

Correlation between the inhibition of phosphorylation and improvement of insulin sensitivity by anti-diabetic PPARγ ligands a , Glucose-tolerance tests in 16-week HFD mice treated with vehicle, rosiglitazone or MRL24 (n=10). b , phosphorylation of PPARγ in WAT. c , The expression of gene sets regulated by PPARγ phosphorylation in WAT (Error bars are s.e.m.; * p
Figure Legend Snippet: Correlation between the inhibition of phosphorylation and improvement of insulin sensitivity by anti-diabetic PPARγ ligands a , Glucose-tolerance tests in 16-week HFD mice treated with vehicle, rosiglitazone or MRL24 (n=10). b , phosphorylation of PPARγ in WAT. c , The expression of gene sets regulated by PPARγ phosphorylation in WAT (Error bars are s.e.m.; * p

Techniques Used: Inhibition, Mouse Assay, Expressing

Specific fat cell gene dysregulation by the cdk5-mediated S273 phosphorylation of PPARγ a , In vitro CDK assays performed using cdk5/p35 with either wild type (WT) or S273A mutated PPARγ. b , Phosphorylation of PPARγ in differentiated 3T3-L1 adipocytes stimulated with TNF-α for the indicated times. c , Phosphorylation of PPARγ in cells expressing scrambled or CDK5 shRNA stimulated with indicated cytokines. NT, no treatment. d , Staining of PPARγ-null fibroblasts expressing WT or S273A mutant PPARγ with Oil-Red-O. e , Gene expression in these cells was analyzed by real-time quantitative PCR (qPCR) for expression of various genes (n=3). f , mRNA expression in transplanted fat pads was analyzed by qPCR (n=5) (Error bars are s.e.m.; * p
Figure Legend Snippet: Specific fat cell gene dysregulation by the cdk5-mediated S273 phosphorylation of PPARγ a , In vitro CDK assays performed using cdk5/p35 with either wild type (WT) or S273A mutated PPARγ. b , Phosphorylation of PPARγ in differentiated 3T3-L1 adipocytes stimulated with TNF-α for the indicated times. c , Phosphorylation of PPARγ in cells expressing scrambled or CDK5 shRNA stimulated with indicated cytokines. NT, no treatment. d , Staining of PPARγ-null fibroblasts expressing WT or S273A mutant PPARγ with Oil-Red-O. e , Gene expression in these cells was analyzed by real-time quantitative PCR (qPCR) for expression of various genes (n=3). f , mRNA expression in transplanted fat pads was analyzed by qPCR (n=5) (Error bars are s.e.m.; * p

Techniques Used: In Vitro, Expressing, shRNA, Staining, Mutagenesis, Real-time Polymerase Chain Reaction

Anti-diabetic PPARγ ligands block CDK5-mediated phosphorylation of PPARγ a , TNF-α-induced phosphorylation of PPARγ in 3T3-L1 adipocytes expressing either WT or Q286P mutant of PPARγ treated with rosiglitazone and/or GW9662. b and c , In vitro CDK5 assay with either rosiglitazone or MRL24. d , Transcriptional activity of a PPAR-derived reporter gene in response to rosiglitazone or MRL24 (n=3). e , Microarray analysis of differentiated PPARγ-null fibroblasts expressing WT (NT, rosiglitazone or MRL24 treated) or S273A mutant PPARγ. f , mRNA expression of genes regulated by the phosphorylation of PPARγ in epididymal fat tissue of mice on either chow or HFD for 13 weeks (n=5). Error bars are s.e.m.; * p
Figure Legend Snippet: Anti-diabetic PPARγ ligands block CDK5-mediated phosphorylation of PPARγ a , TNF-α-induced phosphorylation of PPARγ in 3T3-L1 adipocytes expressing either WT or Q286P mutant of PPARγ treated with rosiglitazone and/or GW9662. b and c , In vitro CDK5 assay with either rosiglitazone or MRL24. d , Transcriptional activity of a PPAR-derived reporter gene in response to rosiglitazone or MRL24 (n=3). e , Microarray analysis of differentiated PPARγ-null fibroblasts expressing WT (NT, rosiglitazone or MRL24 treated) or S273A mutant PPARγ. f , mRNA expression of genes regulated by the phosphorylation of PPARγ in epididymal fat tissue of mice on either chow or HFD for 13 weeks (n=5). Error bars are s.e.m.; * p

Techniques Used: Blocking Assay, Expressing, Mutagenesis, In Vitro, Activity Assay, Derivative Assay, Microarray, Mouse Assay

Differential HDX MS data for PPARγ-LBD ± rosiglitazone and MRL24 a , Histograms showing the percent reduction in HDX for Helix 3 (IRIFQGCQF), the β-sheet region (ISEGQGFMTRE), Helix 12 (QEIYKDLY) and the Helix 2-2’ link region containing the site of CDK5 phosphorylation (KTTDKSPFVIYDM). Values are calculated relative to the measured %D value for apo PPARγ-LBD (n=4; error bars are s.e.m.; ** p
Figure Legend Snippet: Differential HDX MS data for PPARγ-LBD ± rosiglitazone and MRL24 a , Histograms showing the percent reduction in HDX for Helix 3 (IRIFQGCQF), the β-sheet region (ISEGQGFMTRE), Helix 12 (QEIYKDLY) and the Helix 2-2’ link region containing the site of CDK5 phosphorylation (KTTDKSPFVIYDM). Values are calculated relative to the measured %D value for apo PPARγ-LBD (n=4; error bars are s.e.m.; ** p

Techniques Used: Mass Spectrometry

31) Product Images from "TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis"

Article Title: TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis

Journal: Oncogene

doi: 10.1038/s41388-017-0043-9

DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P
Figure Legend Snippet: DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P

Techniques Used: In Vivo, Stable Transfection, Expressing, shRNA, Sequencing, Mouse Assay, Injection

DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.
Figure Legend Snippet: DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.

Techniques Used: Transduction, Cell Culture, Incubation, Recombinant, SDS Page, Staining

32) Product Images from "Integrative discovery of treatments for high-risk neuroblastoma"

Article Title: Integrative discovery of treatments for high-risk neuroblastoma

Journal: Nature Communications

doi: 10.1038/s41467-019-13817-8

GW405833 reduces neuroblastoma growth in vivo. a Mice were engrafted with \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$15 \ \times 1{0}^{6}$$\end{document} 15 × 1 0 6 SK-N-BE(2) cells s.c. and randomized to receive a daily i.p. injection of GW (45 mg/kg; n = 10) or vehicle ( n = 12) for 8 days, starting at the appearance of palpable tumours of \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${ > }0.2\,{\mathrm{cm}}^{3}$$\end{document} > 0.2 cm 3 . b GW405833 significantly impaired the growth of established human tumours (hierarchical linear model). c Point comparison of day 8 tumour volume. d Tumour volume increase from day 0 to day 8. e Post-mortem tumour weight after 8 days of treatment. f Cell proliferation marker MKI67, counted using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 4). g Apoptosis marker cleaved PARP, counted in using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 3). h Representative images of tumour histology (HE), MKI67 and c-PARP localization, bar = 100 μm. Statistics: b Mean, 95% confidence interval, p -value computed from a mixed effects model and corrected for multiple testing using bonferroni correction. c – g Mean, standard deviation, Student’s t -test.
Figure Legend Snippet: GW405833 reduces neuroblastoma growth in vivo. a Mice were engrafted with \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$15 \ \times 1{0}^{6}$$\end{document} 15 × 1 0 6 SK-N-BE(2) cells s.c. and randomized to receive a daily i.p. injection of GW (45 mg/kg; n = 10) or vehicle ( n = 12) for 8 days, starting at the appearance of palpable tumours of \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${ > }0.2\,{\mathrm{cm}}^{3}$$\end{document} > 0.2 cm 3 . b GW405833 significantly impaired the growth of established human tumours (hierarchical linear model). c Point comparison of day 8 tumour volume. d Tumour volume increase from day 0 to day 8. e Post-mortem tumour weight after 8 days of treatment. f Cell proliferation marker MKI67, counted using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 4). g Apoptosis marker cleaved PARP, counted in using ImmunoRatio plugin for ImageJ from 10 to 15 representative fields per specimen (DMSO, GW405833, n = 5; AS601245, n = 3). h Representative images of tumour histology (HE), MKI67 and c-PARP localization, bar = 100 μm. Statistics: b Mean, 95% confidence interval, p -value computed from a mixed effects model and corrected for multiple testing using bonferroni correction. c – g Mean, standard deviation, Student’s t -test.

Techniques Used: In Vivo, Mouse Assay, Injection, Marker, Standard Deviation

33) Product Images from "Chemoprevention of Head and Neck Cancer by Simultaneous Blocking of Epidermal Growth Factor Receptor and Cyclooxygenase-2 Signaling Pathways: Preclinical and Clinical Studies"

Article Title: Chemoprevention of Head and Neck Cancer by Simultaneous Blocking of Epidermal Growth Factor Receptor and Cyclooxygenase-2 Signaling Pathways: Preclinical and Clinical Studies

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

doi: 10.1158/1078-0432.CCR-12-3149

Effect of erlotinib and celecoxib on Tu212 xenograft tumor growth Four groups of mice were orally gavaged with control (0.1% Tween 80 and 0.5% methylcellulose), erlotinib (75 mg/kg), celecoxib (50mg/kg), or the combination (n=8) of erlotinb (75 mg/kg) and celecoxib (50 mg/kg) for 6 days prior to a subcutaneous inoculation of 2 × 10 6 Tu212 cells. The animals were continuously gavaged with the agents 5 days a week for a total of 4 weeks. (A) Tumor volume was measured at the indicated time points. (B) Immunohistochemistry analyses shown as representative H E staining (× 200) were performed for expression of proliferation marker Ki-67, mTOR substrate p-S6, and endothelium marker CD34. (C) Quantification of these biomarkers. * indicates statistical significance (p
Figure Legend Snippet: Effect of erlotinib and celecoxib on Tu212 xenograft tumor growth Four groups of mice were orally gavaged with control (0.1% Tween 80 and 0.5% methylcellulose), erlotinib (75 mg/kg), celecoxib (50mg/kg), or the combination (n=8) of erlotinb (75 mg/kg) and celecoxib (50 mg/kg) for 6 days prior to a subcutaneous inoculation of 2 × 10 6 Tu212 cells. The animals were continuously gavaged with the agents 5 days a week for a total of 4 weeks. (A) Tumor volume was measured at the indicated time points. (B) Immunohistochemistry analyses shown as representative H E staining (× 200) were performed for expression of proliferation marker Ki-67, mTOR substrate p-S6, and endothelium marker CD34. (C) Quantification of these biomarkers. * indicates statistical significance (p

Techniques Used: Mouse Assay, Immunohistochemistry, Staining, Expressing, Marker

Induction of G0/G1 arrest by erlotinib and celecoxib in SCCHN cells SCCHN cell lines Tu212 (A) and Tu686 (B) were treated with erlotinib (1 μM), celecoxib (10 μM), and their combination for 24, 48, and 72 hours. Cell cycle analysis was performed by flow cytometry as described in the Methods section. The average percentages of the cell population arrested at G0/G1 at each time point are presented with standard deviation from three repeated experiments. Significant differences ( p ≤ 0.05) in comparison of the combination treatment with the control and each of the single treatments at all time points are shown. (C) Western blot analyses of cell cycle regulatory proteins in both Tu212 and Tu686 cells treated for 24, 48, and 72 hours. The experiments were repeated three times.
Figure Legend Snippet: Induction of G0/G1 arrest by erlotinib and celecoxib in SCCHN cells SCCHN cell lines Tu212 (A) and Tu686 (B) were treated with erlotinib (1 μM), celecoxib (10 μM), and their combination for 24, 48, and 72 hours. Cell cycle analysis was performed by flow cytometry as described in the Methods section. The average percentages of the cell population arrested at G0/G1 at each time point are presented with standard deviation from three repeated experiments. Significant differences ( p ≤ 0.05) in comparison of the combination treatment with the control and each of the single treatments at all time points are shown. (C) Western blot analyses of cell cycle regulatory proteins in both Tu212 and Tu686 cells treated for 24, 48, and 72 hours. The experiments were repeated three times.

Techniques Used: Cell Cycle Assay, Flow Cytometry, Cytometry, Standard Deviation, Western Blot

Signal transduction pathways affected by erlotinib and celecoxib in SCCHN cells Cell lysates were collected from SCCHN cell lines Tu212 and Tu686 after treatment with erlotinib (E: 1 μM), celecoxib (C: 10 μM), and their combination (EC) for 24, 48, and 72 hours. Untreated cells (NT) were used as a control at each time point. Western blot analyses were performed on total protein extracts from each of the time points to detect the expression levels of proteins involved in EGFR, AKT, mTOR, and COX-2 pathways. β-actin served as a loading control.
Figure Legend Snippet: Signal transduction pathways affected by erlotinib and celecoxib in SCCHN cells Cell lysates were collected from SCCHN cell lines Tu212 and Tu686 after treatment with erlotinib (E: 1 μM), celecoxib (C: 10 μM), and their combination (EC) for 24, 48, and 72 hours. Untreated cells (NT) were used as a control at each time point. Western blot analyses were performed on total protein extracts from each of the time points to detect the expression levels of proteins involved in EGFR, AKT, mTOR, and COX-2 pathways. β-actin served as a loading control.

Techniques Used: Transduction, Western Blot, Expressing

Effects of erlotinib and celecoxib on growth of SCCHN cell lines SCCHN cell lines Tu212 (A) and Tu686 (B) were treated with 1:2 serial dilutions of erlotinib (0-40 μM) and celecoxib (0-20 μM) as single agents and in combination as described in the Methods section. After incubation for 72 hours, SRB assay was used to determine the percentage of survival relative to the untreated cells. CIs at effective doses which resulted in 50% (ED50), 75% (ED75), and 90% (ED90) inhibitory rates (1 - survival rate) were calculated using CalcuSyn software. A CI value of > 1 is antagonism, = 1 is additivity, and
Figure Legend Snippet: Effects of erlotinib and celecoxib on growth of SCCHN cell lines SCCHN cell lines Tu212 (A) and Tu686 (B) were treated with 1:2 serial dilutions of erlotinib (0-40 μM) and celecoxib (0-20 μM) as single agents and in combination as described in the Methods section. After incubation for 72 hours, SRB assay was used to determine the percentage of survival relative to the untreated cells. CIs at effective doses which resulted in 50% (ED50), 75% (ED75), and 90% (ED90) inhibitory rates (1 - survival rate) were calculated using CalcuSyn software. A CI value of > 1 is antagonism, = 1 is additivity, and

Techniques Used: Incubation, Sulforhodamine B Assay, Software

34) Product Images from "A Notch1 Ectodomain Construct Inhibits Endothelial Notch Signaling, Tumor Growth, and Angiogenesis"

Article Title: A Notch1 Ectodomain Construct Inhibits Endothelial Notch Signaling, Tumor Growth, and Angiogenesis

Journal: Cancer research

doi: 10.1158/0008-5472.CAN-07-6499

Notch1 decoy expression disrupts angiogenesis and impairs tumor viability in human NGP xenografts. We have previously reported that human neuroblastoma xenografts in mice have a mature, hierarchical vasculature that is relatively resistant to VEGF blockade
Figure Legend Snippet: Notch1 decoy expression disrupts angiogenesis and impairs tumor viability in human NGP xenografts. We have previously reported that human neuroblastoma xenografts in mice have a mature, hierarchical vasculature that is relatively resistant to VEGF blockade

Techniques Used: Expressing, Mouse Assay

35) Product Images from "MicroRNA-339-5p inhibits colorectal tumorigenesis through regulation of the MDM2/p53 signaling"

Article Title: MicroRNA-339-5p inhibits colorectal tumorigenesis through regulation of the MDM2/p53 signaling

Journal: Oncotarget

doi:

MiR-339-5p increases p53 protein accumulation and its transcriptional activity in response to stress by negatively regulating MDM2 in human colorectal cancer cells (A) MiR-339-5p decreased MDM2 protein levels and increased the p53 protein accumulation and transcriptional activity toward p21 in response to stress in HCT116 cells. (B) MiR-339-5p increased the p53 transcriptional activity toward p21, Puma and Fas in response to stress in HCT116 cells. In A and B: HCT116 p53+/+ and p53−/− cells were transfected with miR-339-5p mimic or miR-con. At 24 h after transfection, cells were treated with 5-FU (50 μM) for 8 h, and analyzed by western-blot (A) and Taqman real-time PCR (B), respectively. The mRNA levels of all genes were normalized to actin. The mRNA levels of genes in untreated cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). (C) MiR-339-5p decreased MDM2 protein levels and increased p53, p21 protein levels in response to stress in RKO cells. RKO p53+/+ and RKO p53−/− cells were treated with 5-FU and analyzed as described in A.
Figure Legend Snippet: MiR-339-5p increases p53 protein accumulation and its transcriptional activity in response to stress by negatively regulating MDM2 in human colorectal cancer cells (A) MiR-339-5p decreased MDM2 protein levels and increased the p53 protein accumulation and transcriptional activity toward p21 in response to stress in HCT116 cells. (B) MiR-339-5p increased the p53 transcriptional activity toward p21, Puma and Fas in response to stress in HCT116 cells. In A and B: HCT116 p53+/+ and p53−/− cells were transfected with miR-339-5p mimic or miR-con. At 24 h after transfection, cells were treated with 5-FU (50 μM) for 8 h, and analyzed by western-blot (A) and Taqman real-time PCR (B), respectively. The mRNA levels of all genes were normalized to actin. The mRNA levels of genes in untreated cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). (C) MiR-339-5p decreased MDM2 protein levels and increased p53, p21 protein levels in response to stress in RKO cells. RKO p53+/+ and RKO p53−/− cells were treated with 5-FU and analyzed as described in A.

Techniques Used: Activity Assay, Transfection, Western Blot, Real-time Polymerase Chain Reaction

MiR-339-5p inhibits the growth of HCT116 xenograft tumors in a largely p53-dependent manner (A, B) MiR-339-5p inhibited the growth of HCT116 xenograft tumors in nude mice in a largely p53-dependent manner. Xenograft tumors were established by s.c. injection of HCT116 p53+/+ and HCT116 p53−/− cells into nude mice. When tumor volumes reached ~60mm 3 , tumors were injected with miR-339-5p mimic or miR-con once every two days for 6 times. (A) Representative tumors were photographed at 12 days after the first treatment with miR-339-5p mimic or miR-con. Scale bar: 10 mm. (B) Upper panels: The growth curves of HCT116 p53+/+ and p53−/− tumors after miR-339-5p injection. The relative volumes of the tumors before treatment at day 0 were designated as 1. Lower panel: The fold reduction of tumor volumes by miR-339-5p injection in both HCT116 p53+/+ and p53−/− tumors. Data are presented as mean ± SD (n=12 for each group). *: p
Figure Legend Snippet: MiR-339-5p inhibits the growth of HCT116 xenograft tumors in a largely p53-dependent manner (A, B) MiR-339-5p inhibited the growth of HCT116 xenograft tumors in nude mice in a largely p53-dependent manner. Xenograft tumors were established by s.c. injection of HCT116 p53+/+ and HCT116 p53−/− cells into nude mice. When tumor volumes reached ~60mm 3 , tumors were injected with miR-339-5p mimic or miR-con once every two days for 6 times. (A) Representative tumors were photographed at 12 days after the first treatment with miR-339-5p mimic or miR-con. Scale bar: 10 mm. (B) Upper panels: The growth curves of HCT116 p53+/+ and p53−/− tumors after miR-339-5p injection. The relative volumes of the tumors before treatment at day 0 were designated as 1. Lower panel: The fold reduction of tumor volumes by miR-339-5p injection in both HCT116 p53+/+ and p53−/− tumors. Data are presented as mean ± SD (n=12 for each group). *: p

Techniques Used: Mouse Assay, Injection

MiR-339-5p negatively regulates MDM2 levels in human colorectal cells through binding to human MDM2 3′-UTR (A) MiR-339-5p decreased MDM2 protein levels and increased p53 protein levels in human colorectal cancer HCT116 and RKO cells. HCT116 p53+/+, HCT116 p53−/−, RKO p53+/+ and RKO p53−/− cells were transfected with miR-339-5p mimic or scrambled miRNA control (miR-con), and the MDM2 and p53 protein levels were measured at 24 h after transfection by western-blot assays. (B) MiR-339-5p decreased MDM2 mRNA levels in HCT116 and RKO cells. The MDM2 mRNA levels were measured by Taqman real-time PCR in cells transfected with miR-339-5p mimic or miR-con, and normalized with actin. The levels of the MDM2 mRNA in control cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). #: p
Figure Legend Snippet: MiR-339-5p negatively regulates MDM2 levels in human colorectal cells through binding to human MDM2 3′-UTR (A) MiR-339-5p decreased MDM2 protein levels and increased p53 protein levels in human colorectal cancer HCT116 and RKO cells. HCT116 p53+/+, HCT116 p53−/−, RKO p53+/+ and RKO p53−/− cells were transfected with miR-339-5p mimic or scrambled miRNA control (miR-con), and the MDM2 and p53 protein levels were measured at 24 h after transfection by western-blot assays. (B) MiR-339-5p decreased MDM2 mRNA levels in HCT116 and RKO cells. The MDM2 mRNA levels were measured by Taqman real-time PCR in cells transfected with miR-339-5p mimic or miR-con, and normalized with actin. The levels of the MDM2 mRNA in control cells transfected with miR-con were designated as 1. Data are presented as mean ± SD (n=3). #: p

Techniques Used: Binding Assay, Transfection, Western Blot, Real-time Polymerase Chain Reaction

MiR-339-5p enhances p53-mediated apoptosis and senescence in response to stress (A) MiR-339-5p enhanced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 5-FU (300 μM), and apoptosis were measured by Annexin V staining in a flow cytometer at 24 or 36 h after treatment. (B) The miR-339-5p inhibitor reduced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with 5-FU and analyzed as described in A. (C) MiR-339-5p enhanced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 100 nM Doxorubicin for 3 days before cellular senescence was measured by SA-β-gal staining. (D) The miR-339-5p inhibitor reduced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with Doxorubicin and analyzed as described in C. In C and D: The upper panels are represented images of SA-β-gal staining. In A-D: Data are presented as mean ± SD (n = 3). #: p
Figure Legend Snippet: MiR-339-5p enhances p53-mediated apoptosis and senescence in response to stress (A) MiR-339-5p enhanced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 5-FU (300 μM), and apoptosis were measured by Annexin V staining in a flow cytometer at 24 or 36 h after treatment. (B) The miR-339-5p inhibitor reduced p53-mediated apoptosis in HCT116 cells treated with 5-FU. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with 5-FU and analyzed as described in A. (C) MiR-339-5p enhanced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were treated with 100 nM Doxorubicin for 3 days before cellular senescence was measured by SA-β-gal staining. (D) The miR-339-5p inhibitor reduced p53-mediated senescence in HCT116 cells treated with Doxorubicin. HCT116 p53+/+ and p53−/− cells transfected with the miR-339-5p inhibitor or miR-con inhibitor were treated with Doxorubicin and analyzed as described in C. In C and D: The upper panels are represented images of SA-β-gal staining. In A-D: Data are presented as mean ± SD (n = 3). #: p

Techniques Used: Transfection, Staining, Flow Cytometry, Cytometry

MiR-339-5p inhibits the migration and invasion of colorectal cancer cells in a largely p53-dependent manner (A) MiR-339-5p inhibited the migration of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into chambers for migration assays. Left panels: Representative images of migrated cells; Right panel: Quantifications of average number of migrated cells per field. (B) MiR-339-5p inhibited the invasion of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into matrigel-coated chambers for invasion assays. Left panels: Representative images of invading cells; Right panel: Quantifications of average number of invading cells per field. (C, D) The miR-339-5p inhibitor promoted the migration (C) and invasion (D) of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p inhibitor or miR-con inhibitor were used for migration and invasion assays as described in A and B, respectively. Data are presented as mean ± SD (n=3). #: p
Figure Legend Snippet: MiR-339-5p inhibits the migration and invasion of colorectal cancer cells in a largely p53-dependent manner (A) MiR-339-5p inhibited the migration of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into chambers for migration assays. Left panels: Representative images of migrated cells; Right panel: Quantifications of average number of migrated cells per field. (B) MiR-339-5p inhibited the invasion of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p mimic or miR-con were seeded into matrigel-coated chambers for invasion assays. Left panels: Representative images of invading cells; Right panel: Quantifications of average number of invading cells per field. (C, D) The miR-339-5p inhibitor promoted the migration (C) and invasion (D) of colorectal cancer cells in a largely p53-dependent manner. HCT116 p53+/+ and p53−/− cells transfected with miR-339-5p inhibitor or miR-con inhibitor were used for migration and invasion assays as described in A and B, respectively. Data are presented as mean ± SD (n=3). #: p

Techniques Used: Migration, Transfection

36) Product Images from "TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis"

Article Title: TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis

Journal: Oncogene

doi: 10.1038/s41388-017-0043-9

DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P
Figure Legend Snippet: DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P

Techniques Used: In Vivo, Stable Transfection, Expressing, shRNA, Sequencing, Mouse Assay, Injection

DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.
Figure Legend Snippet: DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.

Techniques Used: Transduction, Cell Culture, Incubation, Recombinant, SDS Page, Staining

37) Product Images from "Validation of the Hsp70-Bag3 Protein-Protein Interaction as a Potential Therapeutic Target in Cancer"

Article Title: Validation of the Hsp70-Bag3 Protein-Protein Interaction as a Potential Therapeutic Target in Cancer

Journal: Molecular cancer therapeutics

doi: 10.1158/1535-7163.MCT-14-0650

JG-98 is active in MCF7 and HeLa xenograft models. JG-98 treatment of mice with either (A) MCF7 cells or (B) HeLa cells xenografted. See the materials and methods for experimental details. Arrows indicate treatments with JG-98. (C) Treated samples from
Figure Legend Snippet: JG-98 is active in MCF7 and HeLa xenograft models. JG-98 treatment of mice with either (A) MCF7 cells or (B) HeLa cells xenografted. See the materials and methods for experimental details. Arrows indicate treatments with JG-98. (C) Treated samples from

Techniques Used: Mouse Assay

38) Product Images from "Beneficial effects of combining nilotinib and imatinib in preclinical models of BCR-ABL+ leukemias"

Article Title: Beneficial effects of combining nilotinib and imatinib in preclinical models of BCR-ABL+ leukemias

Journal:

doi: 10.1182/blood-2006-06-026377

Inhibition of cellular tyrosine phosphorylation in BCR-ABL–expressing cells by imatinib and nilotinib, combined . Immunoblot showing inhibitory effects of nilotinib and imatinib (alone and combined) on total cellular tyrosine phosphorylation and
Figure Legend Snippet: Inhibition of cellular tyrosine phosphorylation in BCR-ABL–expressing cells by imatinib and nilotinib, combined . Immunoblot showing inhibitory effects of nilotinib and imatinib (alone and combined) on total cellular tyrosine phosphorylation and

Techniques Used: Inhibition, Expressing

In vivo effects of the combination of nilotinib (20 mg/kg) and imatinib (75 mg/kg) on BCR-ABL–expressing cells in a murine leukemia model. Influence of preirradiation on mouse responsivity to nilotinib. (A) Effects of vehicle, nilotinib alone
Figure Legend Snippet: In vivo effects of the combination of nilotinib (20 mg/kg) and imatinib (75 mg/kg) on BCR-ABL–expressing cells in a murine leukemia model. Influence of preirradiation on mouse responsivity to nilotinib. (A) Effects of vehicle, nilotinib alone

Techniques Used: In Vivo, Expressing

Drug combination studies: imatinib and nilotinib against imatinib-resistant, BCR-ABL–expressing cell lines. Proliferation studies showing 3-day treatments of (A) F317L-Ba/F3 cells, (B) M351T-Ba/F3 cells, and (C) F486S-Ba/F3 cells with nilotinib,
Figure Legend Snippet: Drug combination studies: imatinib and nilotinib against imatinib-resistant, BCR-ABL–expressing cell lines. Proliferation studies showing 3-day treatments of (A) F317L-Ba/F3 cells, (B) M351T-Ba/F3 cells, and (C) F486S-Ba/F3 cells with nilotinib,

Techniques Used: Expressing

Induction of apoptosis and inhibition of proliferation of nonmutated BCR-ABL–expressing cells by nilotinib and imatinib. (A) Effects of nilotinib and imatinib, alone and combined, on induction of apoptosis of nonmutated BCR-ABL–expressing
Figure Legend Snippet: Induction of apoptosis and inhibition of proliferation of nonmutated BCR-ABL–expressing cells by nilotinib and imatinib. (A) Effects of nilotinib and imatinib, alone and combined, on induction of apoptosis of nonmutated BCR-ABL–expressing

Techniques Used: Inhibition, Expressing

Induction of apoptosis and inhibition of proliferation of imatinib-resistant BCR-ABL–expressing cells by nilotinib and imatinib. (A) Effects of nilotinib and imatinib, alone and combined, on proliferation (left panel; n = 2) and induction of apoptosis
Figure Legend Snippet: Induction of apoptosis and inhibition of proliferation of imatinib-resistant BCR-ABL–expressing cells by nilotinib and imatinib. (A) Effects of nilotinib and imatinib, alone and combined, on proliferation (left panel; n = 2) and induction of apoptosis

Techniques Used: Inhibition, Expressing

Drug combination studies: imatinib and nilotinib against imatinib-sensitive, BCR-ABL–expressing cell lines
Figure Legend Snippet: Drug combination studies: imatinib and nilotinib against imatinib-sensitive, BCR-ABL–expressing cell lines

Techniques Used: Expressing

In vivo effects of the combination of nilotinib (15-20 mg/kg) and imatinib (50 mg/kg) on BCR-ABL–expressing cells in a murine leukemia model. (A) Effects of vehicle, nilotinib alone (15 mg/kg), imatinib alone (50 mg/kg), or a combination of nilotinib
Figure Legend Snippet: In vivo effects of the combination of nilotinib (15-20 mg/kg) and imatinib (50 mg/kg) on BCR-ABL–expressing cells in a murine leukemia model. (A) Effects of vehicle, nilotinib alone (15 mg/kg), imatinib alone (50 mg/kg), or a combination of nilotinib

Techniques Used: In Vivo, Expressing

Drug combination studies: imatinib and nilotinib against imatinib-sensitive, BCR-ABL–expressing cell lines. Proliferation studies showing 3-day treatments of (A) K562 cells, (B) 32D.p210 cells, and (C) KU812 cells with nilotinib, imatinib, or
Figure Legend Snippet: Drug combination studies: imatinib and nilotinib against imatinib-sensitive, BCR-ABL–expressing cell lines. Proliferation studies showing 3-day treatments of (A) K562 cells, (B) 32D.p210 cells, and (C) KU812 cells with nilotinib, imatinib, or

Techniques Used: Expressing

39) Product Images from "TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis"

Article Title: TWIST1 induces expression of Discoidin Domain Receptor2 (DDR2) to Promote Ovarian Cancer Metastasis

Journal: Oncogene

doi: 10.1038/s41388-017-0043-9

DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P
Figure Legend Snippet: DDR2 influences ovarian cancer cell metastasis in vivo A. A2780 cells stably expressing shRNA targeting sequence for scramble control (shSCRM) or DDR2 (shDDR2). Beta-Actin was used as a protein loading control. B–D. Representative images and quantification of Balb/c Nu mice injected IP with 7.5×106 A2780 shSCRM or shDDR2 cells. Tumor burden was assessed at 14 days post injection in the B entire peritoneal cavity C. Mesentary only D. Omentum only. N=10 mice per group. Means and s.d. Unpaired t-test, *P

Techniques Used: In Vivo, Stable Transfection, Expressing, shRNA, Sequencing, Mouse Assay, Injection

DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.
Figure Legend Snippet: DDR2 influences fibronectin cleavage and spreading by ovarian cancer cells A. ES2 or A2780 shRNAi-depleted of DDR2 or transduced with scrambled control (SCRM) were cultured for 6 hours under serum free conditions on 2mg/mL of collagen I, and cell free media was incubated with recombinant human fibronectin (FN). Extent of cleavage was measured at time intervals of 0.25, 1, and 16 hours by SDS Page followed by coomassie staining. B, C. Quantification of cleavage of intact FN by B. A2780 or C. ES2 plotted as a fraction of remaining intact FN as compared to control. Amount of remaining intact FN (~220 kDa) quantified by densitometry. Data from 3 independent experiments are plotted. D. Quantification of cell area when plated on fibronectin coated hydrogels for 3 hrs. Dots represent individual cell areas, with box plot overlay depicting the median and interquartile ranges of area spread. n > 200 cells quantified for each group. E. Representative images of ES2 shSCRM or shDDR2 cells plated on fibronectin coated hydrogels.

Techniques Used: Transduction, Cell Culture, Incubation, Recombinant, SDS Page, Staining

40) Product Images from "Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice"

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice

Journal: Nature Communications

doi: 10.1038/s41467-018-04982-3

In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P
Figure Legend Snippet: In vivo pharmacokinetics (PK) and antitumor activity. a , b PK of unconjugated N297A anti-HER2 mAb (black cross), VCit (blue circle), SVCit (orange triangle), and EVCit (green square) ADCs ( 3a – c ) in female BALB/c mice ( n = 3). At the indicated time points, blood was collected to quantify total antibody (conjugated and unconjugated, a ) and ADC (conjugated only, b ) by sandwich ELISA. c , d Antitumor activity of anti-HER2 ADCs ( 3a , c ) in the JIMT-1 ( c ) and KPL-4 ( d ) xenograft tumor models (female NCr nude mice, n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models). A single dose of VCit ADC ( 3a , 3 mg kg –1 , blue circle), EVCit ADC ( 3c , 3 mg kg –1 , green square; 1 mg kg –1 , magenta triangle), or vehicle control (gray inversed triangle) was administered to mice when a mean tumor volume reached ~100 mm 3 (indicated with a black arrow). Error bars represent s.e.m. e , f Changes in the percentages of surviving mice over time in the JIMT-1 ( e ) and KPL-4 ( f ) models. The curve of ADC ( 3c ) at 1 mg kg –1 (magenta, e ) is slightly shifted upward for clarity. Mice were euthanized at the pre-defined endpoint (see the Method). * P

Techniques Used: In Vivo, Activity Assay, Mouse Assay, Sandwich ELISA

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Article Snippet: .. Four–seven week old intact male athymic nude mice (nu/nu; NCI, Frederick, MD and NCRNU-M; Taconic, Hudson, NY) were subcutaneously implanted with 2 × 106 cells (1:1, v:v with 50 μl of Matrigel® (Becton Dickson, Franklin Lakes, NJ) in Hank's Buffered Salt Solution; BD). .. After ∼10–14 days, 1 nmol of the PSMA specific fluorescent probe, YC27 [ ] (LI-COR) was injected intraperitoneally (I.P.) and images were acquired at a post-injection (p.i.) time point of 24 hr using a Pearl Impulse Imager (LI-COR).

Article Title: Tea nanoparticle, a safe and biocompatible nanocarrier, greatly potentiates the anticancer activity of doxorubicin
Article Snippet: .. Establishing the tumor xenograft mouse model Male athymic NCR (nu/nu) nude mice (NCRNU M, homozygous, albino; 18–25 g; 6–10 week; Taconic Farms) were used for the tumor xenograft experiments. ..

Article Title: Masitinib Antagonizes ATP-Binding Cassette Subfamily C Member 10-Mediated Paclitaxel Resistance: A Preclinical Study
Article Snippet: .. Male athymic NCR (nu/nu) nude mice (NCRNU M, homozygous, albino; 18–25 g; 10–15 week; Taconic Farms, NY) were used for the tumor-xenograft experiments. ..

Article Title: A brain-penetrant RAF dimer antagonist for the noncanonical BRAF oncoprotein of pediatric low-grade astrocytomas
Article Snippet: .. For the in vivo tumorigenesis study, 2×105 p53 null neuroprogenitor cells bearing BRAFV600E or KIAA1549:BRAF were injected into the right striatum of Icr-SCID or NCr nude mice (Taconic Farms) at the coordinates: A, 0 mm, L, 2.0 mm, and D, 2.0 mm relative to the bregma. ..

Injection:

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice
Article Snippet: .. In vivo xenograft mouse models of human breast cancer To produce KPL-4 or JIMT-1 tumors, female NCr nude mice (6−8 weeks old, Taconic Biosciences) were orthotopically injected into the mammary fat pad with 5−7 × 106 cells suspended in 50 µL of 1:1 PBS/Cultrex® BME Type 3 (Trevigen). .. When the tumor volume reached ~100 mm3 , mice were randomly assigned to three or four groups (n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models) and preconditioned with sterile-filtered human IgG (30 mg kg–1 , Innovative Research, catalog number: IRHUGGFLY1G) injected via the tail vein , .

Article Title: Glutamic acid–valine–citrulline linkers ensure stability and efficacy of antibody–drug conjugates in mice
Article Snippet: .. To produce KPL-4 or JIMT-1 tumors, female NCr nude mice (6−8 weeks old, Taconic Biosciences) were orthotopically injected into the mammary fat pad with 5−7 × 106 cells suspended in 50 µL of 1:1 PBS/Cultrex® BME Type 3 (Trevigen). .. When the tumor volume reached ~100 mm3 , mice were randomly assigned to three or four groups ( n = 3 for vehicle in the KPL-4 model; n = 5 for vehicle in the JIMT-1 model and ADCs in both models) and preconditioned with sterile-filtered human IgG (30 mg kg–1 , Innovative Research, catalog number: IRHUGGFLY1G) injected via the tail vein , .

Article Title: A brain-penetrant RAF dimer antagonist for the noncanonical BRAF oncoprotein of pediatric low-grade astrocytomas
Article Snippet: .. For the in vivo tumorigenesis study, 2×105 p53 null neuroprogenitor cells bearing BRAFV600E or KIAA1549:BRAF were injected into the right striatum of Icr-SCID or NCr nude mice (Taconic Farms) at the coordinates: A, 0 mm, L, 2.0 mm, and D, 2.0 mm relative to the bregma. ..

Imaging:

Article Title: Positron Emission Tomographic Imaging of Copper 64– and Gallium 68–Labeled Chelator Conjugates of the Somatostatin Agonist Tyr3-Octreotate
Article Snippet: .. Imaging studies were performed using NCr nude female mice (6–8 weeks, Taconic Labs) bearing HCT116-SSTR2 tumors either in the shoulder or flank. .. Mice were injected with the 64 Cu- and 68 Ga-labeled Y3-TATE analogues (3.7–8.4 MBq) via the tail vein.

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    Taconic Biosciences p carinii infected athymic mice
    Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. <t>carinii</t> organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P
    P Carinii Infected Athymic Mice, supplied by Taconic Biosciences, used in various techniques. Bioz Stars score: 80/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. <t>carinii</t> organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P
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    Taconic Biosciences 6 wk old ncr nude mice
    Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. <t>carinii</t> organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P
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    Image Search Results


    Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P

    Journal: Infection and Immunity

    Article Title: Urokinase-Type Plasminogen Activator in Inflammatory Cell Recruitment and Host Defense against Pneumocystis carinii in Mice

    doi:

    Figure Lengend Snippet: Inflammatory cell accumulation. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were enumerated 4 weeks after inoculation. Data represent means ± SEM; ∗, P

    Article Snippet: P. carinii -infected athymic mice ( nu/nu on a BALB/c background [Taconic Laboratories, Germantown, N.Y.]) were used as a source of P. carinii in all experiments ( ).

    Techniques: Knock-Out, Mouse Assay

    Histology of P. carinii infection. Lungs of wild-type mice inoculated with P. carinii demonstrated perivascular and peribronchiolar mononuclear cell infiltrates (arrow) (A) but no P. carinii organisms (C). Lungs of uPA knockout mice inoculated with P. carinii demonstrated markedly reduced mononuclear cell inflammation but contained alveolar exudate indicative of P. carinii infection (arrows) (B), confirmed by the presence of P. carinii organisms (arrowheads) (D). (A and B) H E stain; magnification, ×200. (C and D) Gomori methenamine silver stain; magnification, ×400.

    Journal: Infection and Immunity

    Article Title: Urokinase-Type Plasminogen Activator in Inflammatory Cell Recruitment and Host Defense against Pneumocystis carinii in Mice

    doi:

    Figure Lengend Snippet: Histology of P. carinii infection. Lungs of wild-type mice inoculated with P. carinii demonstrated perivascular and peribronchiolar mononuclear cell infiltrates (arrow) (A) but no P. carinii organisms (C). Lungs of uPA knockout mice inoculated with P. carinii demonstrated markedly reduced mononuclear cell inflammation but contained alveolar exudate indicative of P. carinii infection (arrows) (B), confirmed by the presence of P. carinii organisms (arrowheads) (D). (A and B) H E stain; magnification, ×200. (C and D) Gomori methenamine silver stain; magnification, ×400.

    Article Snippet: P. carinii -infected athymic mice ( nu/nu on a BALB/c background [Taconic Laboratories, Germantown, N.Y.]) were used as a source of P. carinii in all experiments ( ).

    Techniques: Infection, Mouse Assay, Knock-Out, Staining, Silver Staining

    Intensity of P. carinii infection. Wild-type ( n = 9) and uPA knockout ( n = 10) mice were inoculated intratracheally with P. carinii organisms, and the intensity of infection was scored 4 weeks after inoculation, using a histologic grading scale. Wild-type mice cleared the P. carinii inoculum, with only a single mouse demonstrating several individual, widely scattered P. carinii cysts. Conversely, uPA knockout mice were uniformly heavily infected with P. carinii , with all mice demonstrating grade 3 or grade 4 infections. Data represent medians obtained from two individual experiments. ∗, P

    Journal: Infection and Immunity

    Article Title: Urokinase-Type Plasminogen Activator in Inflammatory Cell Recruitment and Host Defense against Pneumocystis carinii in Mice

    doi:

    Figure Lengend Snippet: Intensity of P. carinii infection. Wild-type ( n = 9) and uPA knockout ( n = 10) mice were inoculated intratracheally with P. carinii organisms, and the intensity of infection was scored 4 weeks after inoculation, using a histologic grading scale. Wild-type mice cleared the P. carinii inoculum, with only a single mouse demonstrating several individual, widely scattered P. carinii cysts. Conversely, uPA knockout mice were uniformly heavily infected with P. carinii , with all mice demonstrating grade 3 or grade 4 infections. Data represent medians obtained from two individual experiments. ∗, P

    Article Snippet: P. carinii -infected athymic mice ( nu/nu on a BALB/c background [Taconic Laboratories, Germantown, N.Y.]) were used as a source of P. carinii in all experiments ( ).

    Techniques: Infection, Knock-Out, Mouse Assay

    Cellular constituents of BAL fluid. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were characterized 4 weeks after inoculation. Total numbers of lymphocytes (A), macrophages (B), and neutrophils (C) are indicated. Data represent means ± SEM; ∗, P

    Journal: Infection and Immunity

    Article Title: Urokinase-Type Plasminogen Activator in Inflammatory Cell Recruitment and Host Defense against Pneumocystis carinii in Mice

    doi:

    Figure Lengend Snippet: Cellular constituents of BAL fluid. Wild-type ( n = 5) and uPA knockout ( n = 5) mice were inoculated intratracheally with P. carinii organisms, and cells in BAL fluid were characterized 4 weeks after inoculation. Total numbers of lymphocytes (A), macrophages (B), and neutrophils (C) are indicated. Data represent means ± SEM; ∗, P

    Article Snippet: P. carinii -infected athymic mice ( nu/nu on a BALB/c background [Taconic Laboratories, Germantown, N.Y.]) were used as a source of P. carinii in all experiments ( ).

    Techniques: Knock-Out, Mouse Assay