skbr3  (ATCC)


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    Structured Review

    ATCC skbr3
    PUVA therapy inhibits ErbB2 signaling. BT474, <t>SKBR3</t> and MCF7 cells were subjected to the indicated treatment conditions as described in Figure 1 . Western blot analysis was performed on whole cell lysates. Actin steady-state protein levels served as a control to ensure for equal loading of protein. Results are representative of three independent experiments.
    Skbr3, supplied by ATCC, used in various techniques. Bioz Stars score: 97/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Photo-Activated Psoralen Binds the ErbB2 Catalytic Kinase Domain, Blocking ErbB2 Signaling and Triggering Tumor Cell Apoptosis"

    Article Title: Photo-Activated Psoralen Binds the ErbB2 Catalytic Kinase Domain, Blocking ErbB2 Signaling and Triggering Tumor Cell Apoptosis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0088983

    PUVA therapy inhibits ErbB2 signaling. BT474, SKBR3 and MCF7 cells were subjected to the indicated treatment conditions as described in Figure 1 . Western blot analysis was performed on whole cell lysates. Actin steady-state protein levels served as a control to ensure for equal loading of protein. Results are representative of three independent experiments.
    Figure Legend Snippet: PUVA therapy inhibits ErbB2 signaling. BT474, SKBR3 and MCF7 cells were subjected to the indicated treatment conditions as described in Figure 1 . Western blot analysis was performed on whole cell lysates. Actin steady-state protein levels served as a control to ensure for equal loading of protein. Results are representative of three independent experiments.

    Techniques Used: Western Blot

    Inhibition of ErbB2 signaling in response to PUVA is independent of DNA crosslinking. The chemical structures of (A) 8MOP, and (B) 7-methylpyridopsoralen (SMSF032310), which is a derivative of 8MOP that lacks the ability to crosslink DNA, are shown. (C) BT474 and SKBR3 cells were exposed to the indicated treatments. Cell growth and viability assays were performed after 72 hr. P
    Figure Legend Snippet: Inhibition of ErbB2 signaling in response to PUVA is independent of DNA crosslinking. The chemical structures of (A) 8MOP, and (B) 7-methylpyridopsoralen (SMSF032310), which is a derivative of 8MOP that lacks the ability to crosslink DNA, are shown. (C) BT474 and SKBR3 cells were exposed to the indicated treatments. Cell growth and viability assays were performed after 72 hr. P

    Techniques Used: Inhibition

    PUVA antitumor activity in HER2 + breast cancer cells. Cells were pre-treated with the indicated concentrations of 8MOP for 4 hr before UVA irradiation (2J), and then cultured for an additional 72 hr before being analyzed for cell growth (A) BT474; (B) SKBR3; (C) MCF7, (D) HFF and apoptosis (D) BT474; (E) SKBR3. Cells treated with vehicle (0.01% DMSO) alone served as controls. Results represent the mean +/− standard error of triplicate samples, and are representative of three independent experiments.
    Figure Legend Snippet: PUVA antitumor activity in HER2 + breast cancer cells. Cells were pre-treated with the indicated concentrations of 8MOP for 4 hr before UVA irradiation (2J), and then cultured for an additional 72 hr before being analyzed for cell growth (A) BT474; (B) SKBR3; (C) MCF7, (D) HFF and apoptosis (D) BT474; (E) SKBR3. Cells treated with vehicle (0.01% DMSO) alone served as controls. Results represent the mean +/− standard error of triplicate samples, and are representative of three independent experiments.

    Techniques Used: Activity Assay, Irradiation, Cell Culture

    The combination of PUVA with the irreversible pan-ErbB TKI neratinib results in enhanced antitumor activity. The growth and viability of BT474 and SKBR3 cells (top bar graphs) after being subjected to the indicated treatment conditions. The combination of PUVA plus neratinib: P
    Figure Legend Snippet: The combination of PUVA with the irreversible pan-ErbB TKI neratinib results in enhanced antitumor activity. The growth and viability of BT474 and SKBR3 cells (top bar graphs) after being subjected to the indicated treatment conditions. The combination of PUVA plus neratinib: P

    Techniques Used: Activity Assay

    2) Product Images from "FAK inhibition alone or in combination with adjuvant therapies reduces cancer stem cell activity"

    Article Title: FAK inhibition alone or in combination with adjuvant therapies reduces cancer stem cell activity

    Journal: NPJ Breast Cancer

    doi: 10.1038/s41523-021-00263-3

    pTyr397FAK expression is higher in triple negative cell lines and ALDH + cells. FAK inhibition in combination with adjuvant therapies reduces CSC activity. a Representative Western blot demonstrating baseline expression of pTyr397FAK, FAK, and GAPDH in breast cell lines including invasive carcinoma cell lines reflecting all molecular phenotypes. b Illustrative plot of relative density of pTyr397FAK to FAK, with relative density of pTyr397FAK to FAK measured and corrected for GAPDH. pTyr397FAK expression in normal ductal cell line MCF10a used as comparison. One-way ANOVA with post hoc Dunnett’s test ( n = 2 for MCF10a and DCIS.com and n = 3 for IDC cell lines). c Representative Western blot demonstrating pTyr397FAK expression in ALDH + and ALDH − MDA-MB-231 cells with illustrative plot of relative densities shown in ( d ) ( n = 3). e ALDH + cells have increased primary MFE compared to ALDH − expressing cells. Both D E analysed using unpaired two tailed t -test. f FAK inhibition with VS4718 0.5 μM reduced primary MFE as a single agent therapy across all cell lines. This was evaluated alongside 1 μM of Tamoxifen in MCF7, 0.1 μM of Lapatinib in BT474 and SKBr3 cells and 0.1 μM Paclitaxel in MDA-MB-231 and SUM159 cells. Each experiment had a minimum of five biological repeats and six technical replicates. g FAK inhibition reduced mammosphere self-renewal in SKBr3 and SUM159 cells when used as monotherapy and in MDA-MB-231 cells when combined with Paclitaxel. All error bars are mean + SEM. Two-way ANOVA with post hoc Tukey’s test (ns not significant, * p
    Figure Legend Snippet: pTyr397FAK expression is higher in triple negative cell lines and ALDH + cells. FAK inhibition in combination with adjuvant therapies reduces CSC activity. a Representative Western blot demonstrating baseline expression of pTyr397FAK, FAK, and GAPDH in breast cell lines including invasive carcinoma cell lines reflecting all molecular phenotypes. b Illustrative plot of relative density of pTyr397FAK to FAK, with relative density of pTyr397FAK to FAK measured and corrected for GAPDH. pTyr397FAK expression in normal ductal cell line MCF10a used as comparison. One-way ANOVA with post hoc Dunnett’s test ( n = 2 for MCF10a and DCIS.com and n = 3 for IDC cell lines). c Representative Western blot demonstrating pTyr397FAK expression in ALDH + and ALDH − MDA-MB-231 cells with illustrative plot of relative densities shown in ( d ) ( n = 3). e ALDH + cells have increased primary MFE compared to ALDH − expressing cells. Both D E analysed using unpaired two tailed t -test. f FAK inhibition with VS4718 0.5 μM reduced primary MFE as a single agent therapy across all cell lines. This was evaluated alongside 1 μM of Tamoxifen in MCF7, 0.1 μM of Lapatinib in BT474 and SKBr3 cells and 0.1 μM Paclitaxel in MDA-MB-231 and SUM159 cells. Each experiment had a minimum of five biological repeats and six technical replicates. g FAK inhibition reduced mammosphere self-renewal in SKBr3 and SUM159 cells when used as monotherapy and in MDA-MB-231 cells when combined with Paclitaxel. All error bars are mean + SEM. Two-way ANOVA with post hoc Tukey’s test (ns not significant, * p

    Techniques Used: Expressing, Inhibition, Activity Assay, Western Blot, Multiple Displacement Amplification, Two Tailed Test

    3) Product Images from "Bax Translocation Mediated Mitochondrial Apoptosis and Caspase Dependent Photosensitizing Effect of Ficus religiosa on Cancer Cells"

    Article Title: Bax Translocation Mediated Mitochondrial Apoptosis and Caspase Dependent Photosensitizing Effect of Ficus religiosa on Cancer Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0040055

    Effects of FAE on mitochondrial membrane potential and Caspase activation in breast cancer cells and normal cells. (A) The breast cancer cells, MDAMB231, T47D, SKBr3 and normal cells like Human Mammary Epithelial cells and Human Endothelial cells after treatment with FAE at 100 µg/ml for 24 h were stained with 50 nM of TMRM and 0.5 µg/ml of Hoechst 33342 for 15 mins. Then the cells were imaged under fluorescent microscope using DAPI and Rhodamine filter sets using 40× objective. The images were captured with Retiga Exi camera using NIS element software (Nikon). (B) Ac-LEHD-AFC cleavage (Caspase 9 activity) (C) Ac-DEVD-AMC cleavage (Caspase 3/7 activity). MCF-7 cells were treated at IC 50 value of FAE for 24, 36 and 48 h. The results were measured fluorometrically. Values are expressed as mean ± SD of triplicate samples. Significant difference from control value was indicated by *(p
    Figure Legend Snippet: Effects of FAE on mitochondrial membrane potential and Caspase activation in breast cancer cells and normal cells. (A) The breast cancer cells, MDAMB231, T47D, SKBr3 and normal cells like Human Mammary Epithelial cells and Human Endothelial cells after treatment with FAE at 100 µg/ml for 24 h were stained with 50 nM of TMRM and 0.5 µg/ml of Hoechst 33342 for 15 mins. Then the cells were imaged under fluorescent microscope using DAPI and Rhodamine filter sets using 40× objective. The images were captured with Retiga Exi camera using NIS element software (Nikon). (B) Ac-LEHD-AFC cleavage (Caspase 9 activity) (C) Ac-DEVD-AMC cleavage (Caspase 3/7 activity). MCF-7 cells were treated at IC 50 value of FAE for 24, 36 and 48 h. The results were measured fluorometrically. Values are expressed as mean ± SD of triplicate samples. Significant difference from control value was indicated by *(p

    Techniques Used: Activation Assay, Staining, Microscopy, Software, Activity Assay

    4) Product Images from "P-glycoprotein-mediated chemoresistance is reversed by carbonic anhydrase XII inhibitors"

    Article Title: P-glycoprotein-mediated chemoresistance is reversed by carbonic anhydrase XII inhibitors

    Journal: Oncotarget

    doi: 10.18632/oncotarget.13040

    Effects of CAXII inhibitors on intracellular doxorubicin retention in drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicinsensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were grown for 24 h in the presence of 5 μM doxorubicin, alone (–) or in the presence of 5 nM of compounds 1–8. Tariquidar (25 nM; Tar) was included as Pgp inhibitor. The intracellular drug content was measured fluorimetrically. Data are presented as means ± SD ( n = 4). Versus doxorubicin alone (–): *p
    Figure Legend Snippet: Effects of CAXII inhibitors on intracellular doxorubicin retention in drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicinsensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were grown for 24 h in the presence of 5 μM doxorubicin, alone (–) or in the presence of 5 nM of compounds 1–8. Tariquidar (25 nM; Tar) was included as Pgp inhibitor. The intracellular drug content was measured fluorimetrically. Data are presented as means ± SD ( n = 4). Versus doxorubicin alone (–): *p

    Techniques Used: Multiple Displacement Amplification

    Effects of CAXII inhibitors on viability of drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicin-sensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were incubated for 72 h with increasing concentrations (1 nM – 1 mM) of doxorubicin (dox), alone or in the presence of 5 nM of compound 1, 2 and 4, then stained in quadruplicate with neutral red. Tariquidar (25 nM; Tar) was included as a Pgp inhibitor. IC 50 of tariquidar alone was > 10 μM; at 25 nM tariquidar reduced viability ≤ 8.23 ± 1.44 % in each cell line. Data are presented as mean IC 50 ± SD ( n = 4). HT29/DX, A549/DX, U2OS/DX, SaOS/DX versus HT29, A549, U2OS, SaOS cells: * p
    Figure Legend Snippet: Effects of CAXII inhibitors on viability of drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicin-sensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were incubated for 72 h with increasing concentrations (1 nM – 1 mM) of doxorubicin (dox), alone or in the presence of 5 nM of compound 1, 2 and 4, then stained in quadruplicate with neutral red. Tariquidar (25 nM; Tar) was included as a Pgp inhibitor. IC 50 of tariquidar alone was > 10 μM; at 25 nM tariquidar reduced viability ≤ 8.23 ± 1.44 % in each cell line. Data are presented as mean IC 50 ± SD ( n = 4). HT29/DX, A549/DX, U2OS/DX, SaOS/DX versus HT29, A549, U2OS, SaOS cells: * p

    Techniques Used: Multiple Displacement Amplification, Incubation, Staining

    5) Product Images from "Retargeting T Cells for HER2-Positive Tumor Killing by a Bispecific Fv-Fc Antibody"

    Article Title: Retargeting T Cells for HER2-Positive Tumor Killing by a Bispecific Fv-Fc Antibody

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0075589

    FcabCD3 directed killing of tumor cells by T cells at different E:T ratios. Different target cells were incubated with T cells at different E:T ratios. Incubation period for cytotoxicity for all cell lines was 20h except for SKBR3 48h. Cytotoxicity measured during coincubation without the addition of antibodies was subtracted from all samples.
    Figure Legend Snippet: FcabCD3 directed killing of tumor cells by T cells at different E:T ratios. Different target cells were incubated with T cells at different E:T ratios. Incubation period for cytotoxicity for all cell lines was 20h except for SKBR3 48h. Cytotoxicity measured during coincubation without the addition of antibodies was subtracted from all samples.

    Techniques Used: Incubation

    FcabCD3 directed killing of tumor cells by T cells at different bsAb concentrations. Dose-response analysis of serial dilutions of FcabCD3, Fcab and Herceptin on (A) SKBR3, (B) Colo205-luc, (C) MDA-MB-231-luc , (D) K562-luc and B16F10-luc cells at E:T10. Values are mean ±SD of triplicates. Statistical analysis for difference between specific tumor cell lysis was analyzed by one-way ANOVA method using SPSS software (SPSS, p
    Figure Legend Snippet: FcabCD3 directed killing of tumor cells by T cells at different bsAb concentrations. Dose-response analysis of serial dilutions of FcabCD3, Fcab and Herceptin on (A) SKBR3, (B) Colo205-luc, (C) MDA-MB-231-luc , (D) K562-luc and B16F10-luc cells at E:T10. Values are mean ±SD of triplicates. Statistical analysis for difference between specific tumor cell lysis was analyzed by one-way ANOVA method using SPSS software (SPSS, p

    Techniques Used: Multiple Displacement Amplification, Lysis, Software

    FACS analysis of FcabCD3 specificities. As described in Materials and Methods section, SKBR3, Colo205-luc, MDA-MB-231-luc, K562-luc, B16F10-luc cells and T cells were stained with serial dilutions of FcabCD3 and Herceptin. The MFI ratio was calculated by MFI of sample divided by MFI of isotype FITC control by FlowJo software.
    Figure Legend Snippet: FACS analysis of FcabCD3 specificities. As described in Materials and Methods section, SKBR3, Colo205-luc, MDA-MB-231-luc, K562-luc, B16F10-luc cells and T cells were stained with serial dilutions of FcabCD3 and Herceptin. The MFI ratio was calculated by MFI of sample divided by MFI of isotype FITC control by FlowJo software.

    Techniques Used: FACS, Multiple Displacement Amplification, Staining, Software

    6) Product Images from "Improving survival by exploiting tumor dependence on stabilized mutant p53 for treatment"

    Article Title: Improving survival by exploiting tumor dependence on stabilized mutant p53 for treatment

    Journal: Nature

    doi: 10.1038/nature14430

    Ganetespib kills mutp53 human and mouse cells in a mutp53-dependent manner a–c , On a molar basis, ganetespib is > 50-fold more potent than 17AAG in degrading mutp53 and killing human mutp53 cancer cells. MDA468 (p53 R280K) ( a ) and T47D (p53 L194F) ( c ) breast cancer cells, as well as ES2 (p53 S241F) ovarian cancer cells ( b ) were seeded into 6-well plates and treated for 24–48h. After incubation, dead cells were washed off and total protein lysates from only live cells were immunoblotted as indicated. CTB assays on parallel cultures for cell viability show drug activity. ( c ) SkBr3, (p53 R175H) breast cancer cells; DU145, heterozygous (p53 P223L/V274F) prostate cancer cells; MDA231 (R280K) breast cancer cells. Mean ± s.e.m. of four ( b ) or three ( c ) technical replicas, unpaired two-tailed Student’s t -test. p-Akt and p-Erk are also Hsp90 clients; cleaved PARP indicates activated apoptosis. d–f , Ganetespib destabilizes mutp53 but not wtp53 in cultured human ovarian carcinoma cells: EFO21 (p53 C124R) and HOC7 (p53 C275F) ( d ), wtp53 COV434 and COLO704 ( e ), and in human non-small cell lung cancer xenografts H1975 (p53 R273H) ( f ). ( f ) Nude mice bearing tumor xenografts (each lane is an independent tumor) were treated with a single bolus of DMSO or ganetespib (50 mg/kg i.v.). Tumors were harvested at baseline (30 min), 48 hr and 72 hr. Cells were lysed and tumors homogenized and immunoblotted as indicated. Chk1 and CDK1 are other Hsp90 clients, cleaved PARP indicates activated apoptosis. g, h , Ganetespib decreases stabilized mutp53 levels in live Q/− T-lymphoma cells within 24 h, associated with induction of apoptosis. ( g ) Freshly isolated live Q/− T-lymphoma cells were treated with DMSO or 50 nM ganetespib for 24h, followed by immunoblots as indicated. Hsp70 indicates drug activity. Hsc70 is the loading control. ( h ) Death curves of freshly isolated Q/− and p53−/− T-lymphoma cells treated with DMSO or 50 nM ganetespib for the indicated times. CTB and trypan blue exclusion assays are plotted. All values are relative to DMSO treatment at the same time point. Mean ± s.e.m, unpaired two-tailed Student’s t -test, n = 4 independent isolates per genotype for every time point, * p
    Figure Legend Snippet: Ganetespib kills mutp53 human and mouse cells in a mutp53-dependent manner a–c , On a molar basis, ganetespib is > 50-fold more potent than 17AAG in degrading mutp53 and killing human mutp53 cancer cells. MDA468 (p53 R280K) ( a ) and T47D (p53 L194F) ( c ) breast cancer cells, as well as ES2 (p53 S241F) ovarian cancer cells ( b ) were seeded into 6-well plates and treated for 24–48h. After incubation, dead cells were washed off and total protein lysates from only live cells were immunoblotted as indicated. CTB assays on parallel cultures for cell viability show drug activity. ( c ) SkBr3, (p53 R175H) breast cancer cells; DU145, heterozygous (p53 P223L/V274F) prostate cancer cells; MDA231 (R280K) breast cancer cells. Mean ± s.e.m. of four ( b ) or three ( c ) technical replicas, unpaired two-tailed Student’s t -test. p-Akt and p-Erk are also Hsp90 clients; cleaved PARP indicates activated apoptosis. d–f , Ganetespib destabilizes mutp53 but not wtp53 in cultured human ovarian carcinoma cells: EFO21 (p53 C124R) and HOC7 (p53 C275F) ( d ), wtp53 COV434 and COLO704 ( e ), and in human non-small cell lung cancer xenografts H1975 (p53 R273H) ( f ). ( f ) Nude mice bearing tumor xenografts (each lane is an independent tumor) were treated with a single bolus of DMSO or ganetespib (50 mg/kg i.v.). Tumors were harvested at baseline (30 min), 48 hr and 72 hr. Cells were lysed and tumors homogenized and immunoblotted as indicated. Chk1 and CDK1 are other Hsp90 clients, cleaved PARP indicates activated apoptosis. g, h , Ganetespib decreases stabilized mutp53 levels in live Q/− T-lymphoma cells within 24 h, associated with induction of apoptosis. ( g ) Freshly isolated live Q/− T-lymphoma cells were treated with DMSO or 50 nM ganetespib for 24h, followed by immunoblots as indicated. Hsp70 indicates drug activity. Hsc70 is the loading control. ( h ) Death curves of freshly isolated Q/− and p53−/− T-lymphoma cells treated with DMSO or 50 nM ganetespib for the indicated times. CTB and trypan blue exclusion assays are plotted. All values are relative to DMSO treatment at the same time point. Mean ± s.e.m, unpaired two-tailed Student’s t -test, n = 4 independent isolates per genotype for every time point, * p

    Techniques Used: Incubation, CtB Assay, Activity Assay, Two Tailed Test, Cell Culture, Mouse Assay, Isolation, Western Blot

    7) Product Images from "Nuclear factor kappa B activation-induced anti-apoptosis renders HER2 positive cells drug resistant and accelerates tumor growth"

    Article Title: Nuclear factor kappa B activation-induced anti-apoptosis renders HER2 positive cells drug resistant and accelerates tumor growth

    Journal: Molecular cancer research : MCR

    doi: 10.1158/1541-7786.MCR-13-0206-T

    Xenograft growth and apoptotic fractions of SKBR3 and its derivatives. A, Growth of SKBR3 or its HER2-enhanced clonal derivative SKR6. B, SKR6CA. C, SKR6LR. 3×10 6 cells were suspended in 0.25ml PBS mixed with 0.25 ml of matrigel and implanted subcutaneously on the dorsal surface of nu/nu mice. Tumor volume was measured weekly; error bars are 1 standard deviation of the mean of 10 tumors. At 8-10 weeks, tumors from SKR6CA and SKR6LR cells, and at 17 weeks tumors from SKR6 were excised, formalin fixed and paraffin embedded. Sections were stained with Apop Tag™ for apoptotic nuclei. Representative images are shown at 1.5X magnification, and expanded to 20X to visualize the apoptotic fraction. D, Representative section from SKR6 xenografts. E, Representative section from SKR6CA xenografts. F, Representative section from SKR6LR xenografts. G, Mean apoptotic fractions from SKR6, SKR6CA and SKR6LR xenografts were determined by analysis of five independent sections from each of three tumors quantified by image analysis. Error bars are 1 standard deviation of the mean apoptotic fraction.
    Figure Legend Snippet: Xenograft growth and apoptotic fractions of SKBR3 and its derivatives. A, Growth of SKBR3 or its HER2-enhanced clonal derivative SKR6. B, SKR6CA. C, SKR6LR. 3×10 6 cells were suspended in 0.25ml PBS mixed with 0.25 ml of matrigel and implanted subcutaneously on the dorsal surface of nu/nu mice. Tumor volume was measured weekly; error bars are 1 standard deviation of the mean of 10 tumors. At 8-10 weeks, tumors from SKR6CA and SKR6LR cells, and at 17 weeks tumors from SKR6 were excised, formalin fixed and paraffin embedded. Sections were stained with Apop Tag™ for apoptotic nuclei. Representative images are shown at 1.5X magnification, and expanded to 20X to visualize the apoptotic fraction. D, Representative section from SKR6 xenografts. E, Representative section from SKR6CA xenografts. F, Representative section from SKR6LR xenografts. G, Mean apoptotic fractions from SKR6, SKR6CA and SKR6LR xenografts were determined by analysis of five independent sections from each of three tumors quantified by image analysis. Error bars are 1 standard deviation of the mean apoptotic fraction.

    Techniques Used: Mouse Assay, Standard Deviation, Staining

    Functional assessment of NF-κB activation in SKBR3 cells. A, Intracellular localization of NF-κB p65 and HER2 was determined by immunofluorescence. p65 (red, upper left) is found in the nucleus of SKBR3 cells growing exponentially in rich medium. Membranous staining of HER2 (green, upper right) reflects the cell surface localization of the receptor-like protein. Nuclei are stained with DAPI (blue, bottom left). The merged image of p65, HER2 and DAPI is shown (bottom right). B, Cells grown in minimal medium were treated with DMSO (vehicle, top panel), stimulated with 2nM HRG for 18 hours (middle panel), or HRG-stimulated cells were treated with 10μM NBD for 72h. NF-κB p65 is in red and nuclei were stained with DAPI. The inset shows a higher magnification of a single cell. C, The NF-κB DNA binding activity in 10μg of nuclear protein from parental SKBR3 cells cultured in the indicated conditions: Minimal medium stripped of growth factors, rich or complete media, minimal media supplemented with DMSO, minimal media plus 2nM HRG for 18 hours, or HRG plus 10 μg NBD for an additional 72 hours. NF-kB p65 DNA binding was assessed by EMSA. D, DNA Binding activity of indicated genes in SKR6 cells by chromatin immunoprecipitation assay (ChIP). TNF, tumor necrosis factor; TRAF2, TNF-receptor activating Factor 2; NFKB1, nuclear factor kappa B 1; and nuclear factor kappa B 2. E, NF-kB DNA binding activity by EMSA in nuclear proteins. 10μg of nuclear extracts from SKR6 and SKR6 Vector and 5μg of nuclear extracts from SKR6CA and SKR6LR cells were used. Each derivative was grown in rich medium, minimal medium, or minimal medium supplemented as in panel C.
    Figure Legend Snippet: Functional assessment of NF-κB activation in SKBR3 cells. A, Intracellular localization of NF-κB p65 and HER2 was determined by immunofluorescence. p65 (red, upper left) is found in the nucleus of SKBR3 cells growing exponentially in rich medium. Membranous staining of HER2 (green, upper right) reflects the cell surface localization of the receptor-like protein. Nuclei are stained with DAPI (blue, bottom left). The merged image of p65, HER2 and DAPI is shown (bottom right). B, Cells grown in minimal medium were treated with DMSO (vehicle, top panel), stimulated with 2nM HRG for 18 hours (middle panel), or HRG-stimulated cells were treated with 10μM NBD for 72h. NF-κB p65 is in red and nuclei were stained with DAPI. The inset shows a higher magnification of a single cell. C, The NF-κB DNA binding activity in 10μg of nuclear protein from parental SKBR3 cells cultured in the indicated conditions: Minimal medium stripped of growth factors, rich or complete media, minimal media supplemented with DMSO, minimal media plus 2nM HRG for 18 hours, or HRG plus 10 μg NBD for an additional 72 hours. NF-kB p65 DNA binding was assessed by EMSA. D, DNA Binding activity of indicated genes in SKR6 cells by chromatin immunoprecipitation assay (ChIP). TNF, tumor necrosis factor; TRAF2, TNF-receptor activating Factor 2; NFKB1, nuclear factor kappa B 1; and nuclear factor kappa B 2. E, NF-kB DNA binding activity by EMSA in nuclear proteins. 10μg of nuclear extracts from SKR6 and SKR6 Vector and 5μg of nuclear extracts from SKR6CA and SKR6LR cells were used. Each derivative was grown in rich medium, minimal medium, or minimal medium supplemented as in panel C.

    Techniques Used: Functional Assay, Activation Assay, Immunofluorescence, Staining, Binding Assay, Activity Assay, Cell Culture, Chromatin Immunoprecipitation, Plasmid Preparation

    8) Product Images from "Nuclear factor kappa B activation-induced anti-apoptosis renders HER2 positive cells drug resistant and accelerates tumor growth"

    Article Title: Nuclear factor kappa B activation-induced anti-apoptosis renders HER2 positive cells drug resistant and accelerates tumor growth

    Journal: Molecular cancer research : MCR

    doi: 10.1158/1541-7786.MCR-13-0206-T

    Xenograft growth and apoptotic fractions of SKBR3 and its derivatives. A, Growth of SKBR3 or its HER2-enhanced clonal derivative SKR6. B, SKR6CA. C, SKR6LR. 3×10 6 cells were suspended in 0.25ml PBS mixed with 0.25 ml of matrigel and implanted subcutaneously on the dorsal surface of nu/nu mice. Tumor volume was measured weekly; error bars are 1 standard deviation of the mean of 10 tumors. At 8-10 weeks, tumors from SKR6CA and SKR6LR cells, and at 17 weeks tumors from SKR6 were excised, formalin fixed and paraffin embedded. Sections were stained with Apop Tag™ for apoptotic nuclei. Representative images are shown at 1.5X magnification, and expanded to 20X to visualize the apoptotic fraction. D, Representative section from SKR6 xenografts. E, Representative section from SKR6CA xenografts. F, Representative section from SKR6LR xenografts. G, Mean apoptotic fractions from SKR6, SKR6CA and SKR6LR xenografts were determined by analysis of five independent sections from each of three tumors quantified by image analysis. Error bars are 1 standard deviation of the mean apoptotic fraction.
    Figure Legend Snippet: Xenograft growth and apoptotic fractions of SKBR3 and its derivatives. A, Growth of SKBR3 or its HER2-enhanced clonal derivative SKR6. B, SKR6CA. C, SKR6LR. 3×10 6 cells were suspended in 0.25ml PBS mixed with 0.25 ml of matrigel and implanted subcutaneously on the dorsal surface of nu/nu mice. Tumor volume was measured weekly; error bars are 1 standard deviation of the mean of 10 tumors. At 8-10 weeks, tumors from SKR6CA and SKR6LR cells, and at 17 weeks tumors from SKR6 were excised, formalin fixed and paraffin embedded. Sections were stained with Apop Tag™ for apoptotic nuclei. Representative images are shown at 1.5X magnification, and expanded to 20X to visualize the apoptotic fraction. D, Representative section from SKR6 xenografts. E, Representative section from SKR6CA xenografts. F, Representative section from SKR6LR xenografts. G, Mean apoptotic fractions from SKR6, SKR6CA and SKR6LR xenografts were determined by analysis of five independent sections from each of three tumors quantified by image analysis. Error bars are 1 standard deviation of the mean apoptotic fraction.

    Techniques Used: Mouse Assay, Standard Deviation, Staining

    Functional assessment of NF-κB activation in SKBR3 cells. A, Intracellular localization of NF-κB p65 and HER2 was determined by immunofluorescence. p65 (red, upper left) is found in the nucleus of SKBR3 cells growing exponentially in rich medium. Membranous staining of HER2 (green, upper right) reflects the cell surface localization of the receptor-like protein. Nuclei are stained with DAPI (blue, bottom left). The merged image of p65, HER2 and DAPI is shown (bottom right). B, Cells grown in minimal medium were treated with DMSO (vehicle, top panel), stimulated with 2nM HRG for 18 hours (middle panel), or HRG-stimulated cells were treated with 10μM NBD for 72h. NF-κB p65 is in red and nuclei were stained with DAPI. The inset shows a higher magnification of a single cell. C, The NF-κB DNA binding activity in 10μg of nuclear protein from parental SKBR3 cells cultured in the indicated conditions: Minimal medium stripped of growth factors, rich or complete media, minimal media supplemented with DMSO, minimal media plus 2nM HRG for 18 hours, or HRG plus 10 μg NBD for an additional 72 hours. NF-kB p65 DNA binding was assessed by EMSA. D, DNA Binding activity of indicated genes in SKR6 cells by chromatin immunoprecipitation assay (ChIP). TNF, tumor necrosis factor; TRAF2, TNF-receptor activating Factor 2; NFKB1, nuclear factor kappa B 1; and nuclear factor kappa B 2. E, NF-kB DNA binding activity by EMSA in nuclear proteins. 10μg of nuclear extracts from SKR6 and SKR6 Vector and 5μg of nuclear extracts from SKR6CA and SKR6LR cells were used. Each derivative was grown in rich medium, minimal medium, or minimal medium supplemented as in panel C.
    Figure Legend Snippet: Functional assessment of NF-κB activation in SKBR3 cells. A, Intracellular localization of NF-κB p65 and HER2 was determined by immunofluorescence. p65 (red, upper left) is found in the nucleus of SKBR3 cells growing exponentially in rich medium. Membranous staining of HER2 (green, upper right) reflects the cell surface localization of the receptor-like protein. Nuclei are stained with DAPI (blue, bottom left). The merged image of p65, HER2 and DAPI is shown (bottom right). B, Cells grown in minimal medium were treated with DMSO (vehicle, top panel), stimulated with 2nM HRG for 18 hours (middle panel), or HRG-stimulated cells were treated with 10μM NBD for 72h. NF-κB p65 is in red and nuclei were stained with DAPI. The inset shows a higher magnification of a single cell. C, The NF-κB DNA binding activity in 10μg of nuclear protein from parental SKBR3 cells cultured in the indicated conditions: Minimal medium stripped of growth factors, rich or complete media, minimal media supplemented with DMSO, minimal media plus 2nM HRG for 18 hours, or HRG plus 10 μg NBD for an additional 72 hours. NF-kB p65 DNA binding was assessed by EMSA. D, DNA Binding activity of indicated genes in SKR6 cells by chromatin immunoprecipitation assay (ChIP). TNF, tumor necrosis factor; TRAF2, TNF-receptor activating Factor 2; NFKB1, nuclear factor kappa B 1; and nuclear factor kappa B 2. E, NF-kB DNA binding activity by EMSA in nuclear proteins. 10μg of nuclear extracts from SKR6 and SKR6 Vector and 5μg of nuclear extracts from SKR6CA and SKR6LR cells were used. Each derivative was grown in rich medium, minimal medium, or minimal medium supplemented as in panel C.

    Techniques Used: Functional Assay, Activation Assay, Immunofluorescence, Staining, Binding Assay, Activity Assay, Cell Culture, Chromatin Immunoprecipitation, Plasmid Preparation

    9) Product Images from "Histidine-Tagged Folate-Targeted Gold Nanoparticles for Enhanced Transgene Expression in Breast Cancer Cells In Vitro"

    Article Title: Histidine-Tagged Folate-Targeted Gold Nanoparticles for Enhanced Transgene Expression in Breast Cancer Cells In Vitro

    Journal: Pharmaceutics

    doi: 10.3390/pharmaceutics14010053

    Luciferase activity in ( A ) HEK293, ( B ) SKBR3 and ( C ) MCF-7 cells. Data are represented as the mean ± SD ( n = 3). Control 1: untreated cells and Control 2: naked pDNA-treated cells. Lipofectin ® :pDNA complexes at the optimal gene expression were used as a positive control. ** p
    Figure Legend Snippet: Luciferase activity in ( A ) HEK293, ( B ) SKBR3 and ( C ) MCF-7 cells. Data are represented as the mean ± SD ( n = 3). Control 1: untreated cells and Control 2: naked pDNA-treated cells. Lipofectin ® :pDNA complexes at the optimal gene expression were used as a positive control. ** p

    Techniques Used: Luciferase, Activity Assay, Expressing, Positive Control

    Fluorescent images from the acridine orange/ethidium bromide apoptosis assay in the HEK293, SKBR3 and MCF-7 cell lines at 20× magnification. Control = untreated ( A ) HEK293 ( B ) SKBR3 and ( C ) MCF-7 cells. FAuNP treated cells ( D ) HEK293 cells with Au-CS at the suboptimum ratio, ( E ) SKBR3 cells with Au-CS-FA-His at the supraoptimum ratio and ( F ) MCF-7 cells with Au-CS at the optimum ratio. Scale bar = 100 µm.
    Figure Legend Snippet: Fluorescent images from the acridine orange/ethidium bromide apoptosis assay in the HEK293, SKBR3 and MCF-7 cell lines at 20× magnification. Control = untreated ( A ) HEK293 ( B ) SKBR3 and ( C ) MCF-7 cells. FAuNP treated cells ( D ) HEK293 cells with Au-CS at the suboptimum ratio, ( E ) SKBR3 cells with Au-CS-FA-His at the supraoptimum ratio and ( F ) MCF-7 cells with Au-CS at the optimum ratio. Scale bar = 100 µm.

    Techniques Used: Apoptosis Assay

    MTT cytotoxicity assay in the ( A ) HEK293, ( B ) SKBR3 and ( C ) MCF-7 cell lines. Data are represented as the mean ± SD ( n = 3). *** p
    Figure Legend Snippet: MTT cytotoxicity assay in the ( A ) HEK293, ( B ) SKBR3 and ( C ) MCF-7 cell lines. Data are represented as the mean ± SD ( n = 3). *** p

    Techniques Used: MTT Assay, Cytotoxicity Assay

    10) Product Images from "Let-7a regulates mammosphere formation capacity through Ras/NF-κB and Ras/MAPK/ERK pathway in breast cancer stem cells"

    Article Title: Let-7a regulates mammosphere formation capacity through Ras/NF-κB and Ras/MAPK/ERK pathway in breast cancer stem cells

    Journal: Cell Cycle

    doi: 10.1080/15384101.2015.1030547

    Let-7a regulates NF-κB and MAPK/ERK pathway in a KRas-dependent manner in breast cancer cells ( A and B ) The protein expression levels were determined in MCF-7 and SKBR3 cells transfected lenti- let-7a and/or KRas-siRNA by western blot. ( C and
    Figure Legend Snippet: Let-7a regulates NF-κB and MAPK/ERK pathway in a KRas-dependent manner in breast cancer cells ( A and B ) The protein expression levels were determined in MCF-7 and SKBR3 cells transfected lenti- let-7a and/or KRas-siRNA by western blot. ( C and

    Techniques Used: Expressing, Transfection, Western Blot

    KRas is a direct target of let-7a. ( A ) The protein expression levels were determined in MCF-7 and SKBR3 cells transfected lenti-let-7a or lenti-scramble by western blot. ( B-D ) Luciferase activities were analyzed in 2 breast cell lines 36h after cotransfection
    Figure Legend Snippet: KRas is a direct target of let-7a. ( A ) The protein expression levels were determined in MCF-7 and SKBR3 cells transfected lenti-let-7a or lenti-scramble by western blot. ( B-D ) Luciferase activities were analyzed in 2 breast cell lines 36h after cotransfection

    Techniques Used: Expressing, Transfection, Western Blot, Luciferase, Cotransfection

    11) Product Images from "Co-targeting BET bromodomain BRD4 and RAC1 suppresses growth, stemness and tumorigenesis by disrupting the c-MYC-G9a-FTH1axis and downregulating HDAC1 in molecular subtypes of breast cancer"

    Article Title: Co-targeting BET bromodomain BRD4 and RAC1 suppresses growth, stemness and tumorigenesis by disrupting the c-MYC-G9a-FTH1axis and downregulating HDAC1 in molecular subtypes of breast cancer

    Journal: International Journal of Biological Sciences

    doi: 10.7150/ijbs.62236

    Co-targeting BRD4 and RAC1 targets c-MYC-G9a-FTH1 axis and induces autophagy. The expression of c-MYC, G9a and FTH1 was examined in MCF-7, MDA-MB- 231, SKBR3 and JIMT-1 cells treated with ( A ) JQ1 (2 uM) and/or NSC (30 uM) or combination for 72 h; β-actin was used as a negative loading control. Expression status of ( B ) LC3BI/II was evaluated in MCF-7, MDA-MB-231, SKBR3 and/or JIMT-1 cells at 72 h following treatment with JQ1 (2 uM) and/or NSC (30 uM) for 72 h. β-actin was used as a negative loading control.
    Figure Legend Snippet: Co-targeting BRD4 and RAC1 targets c-MYC-G9a-FTH1 axis and induces autophagy. The expression of c-MYC, G9a and FTH1 was examined in MCF-7, MDA-MB- 231, SKBR3 and JIMT-1 cells treated with ( A ) JQ1 (2 uM) and/or NSC (30 uM) or combination for 72 h; β-actin was used as a negative loading control. Expression status of ( B ) LC3BI/II was evaluated in MCF-7, MDA-MB-231, SKBR3 and/or JIMT-1 cells at 72 h following treatment with JQ1 (2 uM) and/or NSC (30 uM) for 72 h. β-actin was used as a negative loading control.

    Techniques Used: Expressing, Multiple Displacement Amplification

    Vitamin C further sensitizes BRCA cells to the anti-growth effects of JQ1 plus NSC treatment. Cell viability as determined by ( A ) MTT assay or ( B ) Crystal violet staining in MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells treated with vitamin C (10 or 20 uM), JQ1 (0.5 uM) and NSC (15 uM) for 72 h. (C) Mean ±SD of crystal violet absorbance values as calculated based on three independent experiments. p
    Figure Legend Snippet: Vitamin C further sensitizes BRCA cells to the anti-growth effects of JQ1 plus NSC treatment. Cell viability as determined by ( A ) MTT assay or ( B ) Crystal violet staining in MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells treated with vitamin C (10 or 20 uM), JQ1 (0.5 uM) and NSC (15 uM) for 72 h. (C) Mean ±SD of crystal violet absorbance values as calculated based on three independent experiments. p

    Techniques Used: MTT Assay, Staining, Multiple Displacement Amplification

    Combined JQ1 plus NSC treatment does not promote ferroptosis in BRCA cells. (A) Expression of SLC7A11 and GPX4 proteins in MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells at 72 h -post-treatment with JQ1 (2 uM) and/or NSC (30 uM) for 72 h; β-actin was used as a negative loading control. (B) Cell viability as determined by crystal violet staining in MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells treated with Ferrostatin-1 (Fer-1) for 72 h in the presence/absence of JQ1 (2 uM) plus NSC (30 uM). (C) Mean ±SD of crystal violet absorbance values as calculated based on three independent experiments. *p
    Figure Legend Snippet: Combined JQ1 plus NSC treatment does not promote ferroptosis in BRCA cells. (A) Expression of SLC7A11 and GPX4 proteins in MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells at 72 h -post-treatment with JQ1 (2 uM) and/or NSC (30 uM) for 72 h; β-actin was used as a negative loading control. (B) Cell viability as determined by crystal violet staining in MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells treated with Ferrostatin-1 (Fer-1) for 72 h in the presence/absence of JQ1 (2 uM) plus NSC (30 uM). (C) Mean ±SD of crystal violet absorbance values as calculated based on three independent experiments. *p

    Techniques Used: Expressing, Multiple Displacement Amplification, Staining

    Combined JQ1 plus NSC treatment suppresses growth and oncogenic potential of different molecular subtypes of BRCA cells. MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with increasing concentrations of JQ1 and/or NSC for 72 h and assessed for viability by ( A ) MTT assay and ( B ) crystal violet staining. (C) Mean ± SD of crystal violet absorbance values based on three independent experiments. Data presented in A and C is the mean ±SD of three independent experiments; *p
    Figure Legend Snippet: Combined JQ1 plus NSC treatment suppresses growth and oncogenic potential of different molecular subtypes of BRCA cells. MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with increasing concentrations of JQ1 and/or NSC for 72 h and assessed for viability by ( A ) MTT assay and ( B ) crystal violet staining. (C) Mean ± SD of crystal violet absorbance values based on three independent experiments. Data presented in A and C is the mean ±SD of three independent experiments; *p

    Techniques Used: Multiple Displacement Amplification, MTT Assay, Staining

    Combined JQ1 plus NSC treatment induces cellular senescence, inhibits mammosphere formation and cell migration in breast cancer cells. (A) MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with JQ1 (0.5 uM) and/or NSC (15 uM) for 72 h and further grown in drug-free medium for 7 days to determine cellular senescence. (B) Statistical analysis of data from A; data presented is the mean ±SD of three independent experiments. *p
    Figure Legend Snippet: Combined JQ1 plus NSC treatment induces cellular senescence, inhibits mammosphere formation and cell migration in breast cancer cells. (A) MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with JQ1 (0.5 uM) and/or NSC (15 uM) for 72 h and further grown in drug-free medium for 7 days to determine cellular senescence. (B) Statistical analysis of data from A; data presented is the mean ±SD of three independent experiments. *p

    Techniques Used: Migration, Multiple Displacement Amplification

    Effects of JQ1 and NSC treatment on the expression of key cellular signaling pathways and HDAC1/Ac-H3K9 axis. (A) The expression of BRD4, RAC1 and NFKB1 were examined in MCF-7, MDA-MB- 231, SKBR3 and JIMT-1 cells treated with JQ1 (2 µM) and/or NSC (30 µM) or combination for 72 h; β-actin was used as a negative loading control. (B) The expression of HDAC1 and AC-H3K9 were determined in MCF-7, MDA-MB- 231, SKBR3 and JIMT-1 cells treated with JQ1 (2 µM) and/or NSC (30 µM) or combination for 72 h; β-actin was used as a negative loading control.
    Figure Legend Snippet: Effects of JQ1 and NSC treatment on the expression of key cellular signaling pathways and HDAC1/Ac-H3K9 axis. (A) The expression of BRD4, RAC1 and NFKB1 were examined in MCF-7, MDA-MB- 231, SKBR3 and JIMT-1 cells treated with JQ1 (2 µM) and/or NSC (30 µM) or combination for 72 h; β-actin was used as a negative loading control. (B) The expression of HDAC1 and AC-H3K9 were determined in MCF-7, MDA-MB- 231, SKBR3 and JIMT-1 cells treated with JQ1 (2 µM) and/or NSC (30 µM) or combination for 72 h; β-actin was used as a negative loading control.

    Techniques Used: Expressing, Multiple Displacement Amplification

    Treatment with JQ1 or NSC inhibits different molecular subtypes of BRCA cell growth. MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with increasing concentrations of JQ1 for 72 h and assessed for viability by ( A ) MTT assay and ( B ) crystal violet staining. (C) Mean ± SD of crystal violet absorbance values based on three independent experiments. MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with increasing concentrations of NSC for 72 h and assessed for viability by ( D ) MTT assay and ( E ) crystal violet staining. (F) Mean ± SD of crystal violet absorbance values based on three independent experiments. Data presented is the mean ± SD of three independent experiments; *p
    Figure Legend Snippet: Treatment with JQ1 or NSC inhibits different molecular subtypes of BRCA cell growth. MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with increasing concentrations of JQ1 for 72 h and assessed for viability by ( A ) MTT assay and ( B ) crystal violet staining. (C) Mean ± SD of crystal violet absorbance values based on three independent experiments. MCF-7, MDA-MB-231, SKBR3 and JIMT-1 cells were treated with increasing concentrations of NSC for 72 h and assessed for viability by ( D ) MTT assay and ( E ) crystal violet staining. (F) Mean ± SD of crystal violet absorbance values based on three independent experiments. Data presented is the mean ± SD of three independent experiments; *p

    Techniques Used: Multiple Displacement Amplification, MTT Assay, Staining

    12) Product Images from "Orlistat and antisense-miRNA-loaded PLGA-PEG nanoparticles for enhanced triple negative breast cancer therapy"

    Article Title: Orlistat and antisense-miRNA-loaded PLGA-PEG nanoparticles for enhanced triple negative breast cancer therapy

    Journal: Nanomedicine

    doi: 10.2217/nnm.15.193

    Cell viability of MDA-MB-231, SKBr3 and MCF10A cells after treated with various concentrations of free orlistat or orlistat nanoparticles ranging from 0 to 10 μM, assessed by MTT assay at 24, 48 and 72 h.
    Figure Legend Snippet: Cell viability of MDA-MB-231, SKBr3 and MCF10A cells after treated with various concentrations of free orlistat or orlistat nanoparticles ranging from 0 to 10 μM, assessed by MTT assay at 24, 48 and 72 h.

    Techniques Used: Multiple Displacement Amplification, MTT Assay

    13) Product Images from "t10c12 Conjugated Linoleic Acid Suppresses HER2 Protein and Enhances Apoptosis in SKBr3 Breast Cancer Cells: Possible Role of COX2"

    Article Title: t10c12 Conjugated Linoleic Acid Suppresses HER2 Protein and Enhances Apoptosis in SKBr3 Breast Cancer Cells: Possible Role of COX2

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0005342

    t10c12 CLA reduces nuclear p65 in SKBr3 cells. (A) Western blots of NFκB p65 in nuclear extracts. Cells were plated in 6-well plates, 3 wells per treatment. Cells from 3 wells were pulled and nuclear extract was obtained as described in Materials and Methods . 10 µg of nuclear extract was loaded into 10% gels. Gel electrophoresis and immunoblots were performed as described in Materials and Methods . Densitometry of bands was performed using Scion Image software Alpha 4.0.3.2. Protein expression of p65 was normalized to total protein and compared to levels in untreated cells (vehicle only) (* = 0.05). Values represent means +/− std error relative to vehicle control from 3 independent experiments. (B) Immunofluorescence of p65. Cells were treated as above. Immunoflourescence was performed as described in Materials and Methods . Top panel: vehicle; Middle pane: 40 µM CLA; Bottom panel: 80 µM CLA. Images were obtained using Delta Vision deconvolution microscope with SoftwoRX 3.5.0 software at 40× and optimized using Adobe PhotoShop CS2 version 9.0.2. (C) Western blot of total and phosphorylated IkappaB protein. Cells were plated and treated as above. 25 µg of whole cell lysate were loaded into 10% gels. Expression levels of IκB proteins were normalized to beta actin and compared to levels in untreated cells (vehicle only). Values represent means +/− std dev from 3 independent experiments (* = 0.05). Gel electrophoresis and immunoblots were performed as described in Materials and Methods . Densitometry of bands was performed using Scion Image software Alpha 4.0.3.2.
    Figure Legend Snippet: t10c12 CLA reduces nuclear p65 in SKBr3 cells. (A) Western blots of NFκB p65 in nuclear extracts. Cells were plated in 6-well plates, 3 wells per treatment. Cells from 3 wells were pulled and nuclear extract was obtained as described in Materials and Methods . 10 µg of nuclear extract was loaded into 10% gels. Gel electrophoresis and immunoblots were performed as described in Materials and Methods . Densitometry of bands was performed using Scion Image software Alpha 4.0.3.2. Protein expression of p65 was normalized to total protein and compared to levels in untreated cells (vehicle only) (* = 0.05). Values represent means +/− std error relative to vehicle control from 3 independent experiments. (B) Immunofluorescence of p65. Cells were treated as above. Immunoflourescence was performed as described in Materials and Methods . Top panel: vehicle; Middle pane: 40 µM CLA; Bottom panel: 80 µM CLA. Images were obtained using Delta Vision deconvolution microscope with SoftwoRX 3.5.0 software at 40× and optimized using Adobe PhotoShop CS2 version 9.0.2. (C) Western blot of total and phosphorylated IkappaB protein. Cells were plated and treated as above. 25 µg of whole cell lysate were loaded into 10% gels. Expression levels of IκB proteins were normalized to beta actin and compared to levels in untreated cells (vehicle only). Values represent means +/− std dev from 3 independent experiments (* = 0.05). Gel electrophoresis and immunoblots were performed as described in Materials and Methods . Densitometry of bands was performed using Scion Image software Alpha 4.0.3.2.

    Techniques Used: Western Blot, Nucleic Acid Electrophoresis, Software, Expressing, Immunofluorescence, Microscopy

    Suppression of HER2 and COX2 in SKBr3 cells. Cells were treated with t10c12 CLA for 24 hrs or Celecoxib for 48 hrs. Cells were plated in 6-well plates, 3 wells per treatment. 30 minutes before collection, 15 µM arachidonic acid was added as substrate for COX2. Negative controls did not receive arachadonic acid. Cells from 3 wells were pulled for isolation of total protein and PGE 2 determination by ELISA. (A) Western blots of HER2 protein following treatment with t10c12 CLA or celecoxib. Gel electrophoresis and immunoblots were performed as described in Materials and Methods . Densitometry of bands was performed using Scion Image software Alpha 4.0.3.2. (B) PGE2 levels were measured by ELISA and are presented as pg/ml relative to vehicle. Values represent the means +/− std dev. from 2 independent experiments. (* = 0.05, ** = 0.001). Experiments were performed as describe in Materials and Methods .
    Figure Legend Snippet: Suppression of HER2 and COX2 in SKBr3 cells. Cells were treated with t10c12 CLA for 24 hrs or Celecoxib for 48 hrs. Cells were plated in 6-well plates, 3 wells per treatment. 30 minutes before collection, 15 µM arachidonic acid was added as substrate for COX2. Negative controls did not receive arachadonic acid. Cells from 3 wells were pulled for isolation of total protein and PGE 2 determination by ELISA. (A) Western blots of HER2 protein following treatment with t10c12 CLA or celecoxib. Gel electrophoresis and immunoblots were performed as described in Materials and Methods . Densitometry of bands was performed using Scion Image software Alpha 4.0.3.2. (B) PGE2 levels were measured by ELISA and are presented as pg/ml relative to vehicle. Values represent the means +/− std dev. from 2 independent experiments. (* = 0.05, ** = 0.001). Experiments were performed as describe in Materials and Methods .

    Techniques Used: Isolation, Enzyme-linked Immunosorbent Assay, Western Blot, Nucleic Acid Electrophoresis, Software

    Proposed mechanism of CLA action in SKBr3 cells. (A) This scenario describes a direct effect on IKK by t10c12 CLA. A consequence of IKK inhibition is reduced phosphorylation of IκB and nuclear localization of p65. A decrease in COX2-derived PGE 2 synthesis will result in loss of HER2 (as described in by Benoit et al, Oncogene, 2004; 23(8):1631). (B) This scenario describes a direct effect on HER2 protein by t10c12 CLA. In this scenario, HER2 is dissociated from the plasma membrane and targeted for ubiquitination and proteosomal degradation. Loss of HER2 signaling downregulates PI-3 kinase and IKK activity. NFκB is sequestered in the cyotosol by IκB and its target genes such as COX2 are not transcribed.
    Figure Legend Snippet: Proposed mechanism of CLA action in SKBr3 cells. (A) This scenario describes a direct effect on IKK by t10c12 CLA. A consequence of IKK inhibition is reduced phosphorylation of IκB and nuclear localization of p65. A decrease in COX2-derived PGE 2 synthesis will result in loss of HER2 (as described in by Benoit et al, Oncogene, 2004; 23(8):1631). (B) This scenario describes a direct effect on HER2 protein by t10c12 CLA. In this scenario, HER2 is dissociated from the plasma membrane and targeted for ubiquitination and proteosomal degradation. Loss of HER2 signaling downregulates PI-3 kinase and IKK activity. NFκB is sequestered in the cyotosol by IκB and its target genes such as COX2 are not transcribed.

    Techniques Used: Inhibition, Derivative Assay, Activity Assay

    14) Product Images from "Down-Regulation of Vitamin D Receptor in Mammospheres: Implications for Vitamin D Resistance in Breast Cancer and Potential for Combination Therapy"

    Article Title: Down-Regulation of Vitamin D Receptor in Mammospheres: Implications for Vitamin D Resistance in Breast Cancer and Potential for Combination Therapy

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0053287

    A, PCR Array Analysis of EMT-specific genes in SKBR3 cells grown under plas and mams conditions after 4 days. B, Validation of PCR Array data by quantitative real time PCR analysis. Experiment was performed in triplicates from three different sets of experiments (*, p≤0.05; **, p≤0.01).
    Figure Legend Snippet: A, PCR Array Analysis of EMT-specific genes in SKBR3 cells grown under plas and mams conditions after 4 days. B, Validation of PCR Array data by quantitative real time PCR analysis. Experiment was performed in triplicates from three different sets of experiments (*, p≤0.05; **, p≤0.01).

    Techniques Used: Polymerase Chain Reaction, Micro-arrays for Mass Spectrometry, Real-time Polymerase Chain Reaction

    A, Analysis of VDR expression in SKBR3 cells grown under mam or plas conditions with or without growth factors (GF: EGF plus FGF). 30 µg of cell lysates were electrophoretically separated and analyzed by western blot analysis using anti-VDR antibody. Membranes were stripped and re-probed using anti-GAPDH antibody for loading controls. B (Left Panel), Analysis of VDR and CD44 protein expression in ALDH − and ALDH + populations isolated from SKBR3 cells. 30 µg of cell lysates obtained from ALDH + and ALDH − cells were electrophoretically separated and analyzed by western blot analysis using anti-VDR or anti-CD44 antibodies. Membranes were subsequently stripped and re-probed with anti-GAPDH antibody for loading controls. B ( Right Panel) , Normalized densitometric ratios showing relative expression levels of VDR and CD44. C (Left Panel), Analysis of VDR and CD44 protein expression in non-transplantable xenografts (NTX) and transplantable xenografts (TX). 30 µg of total cell lysates obtained from non-transplantable xenografts (NTX) and transplantable xenografts (TX) were electrophoretically separated and analyzed by western blot analysis using anti-VDR or anti-CD44 antibodies. Membranes were stripped and re-probed with anti-GAPDH antibody for loading controls (*, p≤0.05; **, p≤0.01). C (Right Panel), Normalized densitometric ratios showing relative expression levels of VDR and CD44.
    Figure Legend Snippet: A, Analysis of VDR expression in SKBR3 cells grown under mam or plas conditions with or without growth factors (GF: EGF plus FGF). 30 µg of cell lysates were electrophoretically separated and analyzed by western blot analysis using anti-VDR antibody. Membranes were stripped and re-probed using anti-GAPDH antibody for loading controls. B (Left Panel), Analysis of VDR and CD44 protein expression in ALDH − and ALDH + populations isolated from SKBR3 cells. 30 µg of cell lysates obtained from ALDH + and ALDH − cells were electrophoretically separated and analyzed by western blot analysis using anti-VDR or anti-CD44 antibodies. Membranes were subsequently stripped and re-probed with anti-GAPDH antibody for loading controls. B ( Right Panel) , Normalized densitometric ratios showing relative expression levels of VDR and CD44. C (Left Panel), Analysis of VDR and CD44 protein expression in non-transplantable xenografts (NTX) and transplantable xenografts (TX). 30 µg of total cell lysates obtained from non-transplantable xenografts (NTX) and transplantable xenografts (TX) were electrophoretically separated and analyzed by western blot analysis using anti-VDR or anti-CD44 antibodies. Membranes were stripped and re-probed with anti-GAPDH antibody for loading controls (*, p≤0.05; **, p≤0.01). C (Right Panel), Normalized densitometric ratios showing relative expression levels of VDR and CD44.

    Techniques Used: Expressing, Western Blot, Isolation

    Selective down-regulation of VDR/RXR expression in mammospheres isolated from breast cancer cells A, Top panel: 50 µg of total cell lysates isolated from breast cancer cell lines SKBR3 (left), MCF7 (middle) and HRas (right) cells grown under plas or mam conditions and were analyzed for VDR and RXR protein expression by Western blot analysis. Bottom panel: Quantitative densitometric analysis of VDR and RXR expression in SKBR3 (left), MCF7 (middle) and HRas (right) cell normalized to GAPDH (*, p≤0.05; **, p≤0.01). B, Top panel: 50 µg of total cell lysates isolated from mammary epithelial HMLE cells grown under plas or mam conditions were analyzed for VDR and RXR protein expression by Western blot analysis. Bottom panel: Quantitative densitometric analysis of VDR and RXR expression normalized to GAPDH (*, p≤0.05; **, p≤0.01). C, Real-time quantitative PCR analysis of VDR expression in SKBR3, MCF7 and HRas as well as in HMLE cells grown under plas or mam conditions (*, p≤0.05; **, p≤0.01). D, Selective up-regulation of Snail in MCF7 and SKBR3 cells. Top panel; 50 µg of total cell lysates isolated from MCF7 (left panel), SKBR3 (middle panel) or HMLE cells (right panel) grown under plas or mam conditions were analyzed for Snail expression by Western blot analysis. Bottom panel: Quantitative densitometric analysis of Snail expression normalized to GAPDH (**, p≤0.01).
    Figure Legend Snippet: Selective down-regulation of VDR/RXR expression in mammospheres isolated from breast cancer cells A, Top panel: 50 µg of total cell lysates isolated from breast cancer cell lines SKBR3 (left), MCF7 (middle) and HRas (right) cells grown under plas or mam conditions and were analyzed for VDR and RXR protein expression by Western blot analysis. Bottom panel: Quantitative densitometric analysis of VDR and RXR expression in SKBR3 (left), MCF7 (middle) and HRas (right) cell normalized to GAPDH (*, p≤0.05; **, p≤0.01). B, Top panel: 50 µg of total cell lysates isolated from mammary epithelial HMLE cells grown under plas or mam conditions were analyzed for VDR and RXR protein expression by Western blot analysis. Bottom panel: Quantitative densitometric analysis of VDR and RXR expression normalized to GAPDH (*, p≤0.05; **, p≤0.01). C, Real-time quantitative PCR analysis of VDR expression in SKBR3, MCF7 and HRas as well as in HMLE cells grown under plas or mam conditions (*, p≤0.05; **, p≤0.01). D, Selective up-regulation of Snail in MCF7 and SKBR3 cells. Top panel; 50 µg of total cell lysates isolated from MCF7 (left panel), SKBR3 (middle panel) or HMLE cells (right panel) grown under plas or mam conditions were analyzed for Snail expression by Western blot analysis. Bottom panel: Quantitative densitometric analysis of Snail expression normalized to GAPDH (**, p≤0.01).

    Techniques Used: Expressing, Isolation, Western Blot, Real-time Polymerase Chain Reaction

    A, Inhibition of VDR expression in SKBR3 cells by siRNA and analysis of VDR, Snail and E-cad protein expression. SKBR3 cells were transfected either with scrambled siRNA (Scram) or On-target smart pool VDR siRNA (siRNA) using standard techniques and protein expression was analyzed by western blot analysis. B, Analysis of EMT-specific gene signature in Scam and VDR siRNA transfected cells (*, p≤0.05; **, p≤0.01). C, Analysis of mammospheres forming capability in Scam and VDR siRNA transfected cells. 5,000 cells after transfection were plated on 24 well ultra-low attachment plates and total numbers of mammospheres formed were analyzed (**, p≤0.01). D, Over-expression of full length human VDR gene and analysis of VDR, Snail and E-cad protein expression by western blot. E, Analysis of EMT-specific gene signature in cells transfected either with control vector or full-length human VDR (*, p≤0.05; **, p≤0.01). F, Analysis of mammospheres forming capability in cells transfected either with control vector or VDR over-expressing (VDR OE) plasmid. 5,000 cells after transfection were plated on 24 well ultra-low attachment plates and total numbers of mammospheres formed were analyzed (**, p≤0.01).
    Figure Legend Snippet: A, Inhibition of VDR expression in SKBR3 cells by siRNA and analysis of VDR, Snail and E-cad protein expression. SKBR3 cells were transfected either with scrambled siRNA (Scram) or On-target smart pool VDR siRNA (siRNA) using standard techniques and protein expression was analyzed by western blot analysis. B, Analysis of EMT-specific gene signature in Scam and VDR siRNA transfected cells (*, p≤0.05; **, p≤0.01). C, Analysis of mammospheres forming capability in Scam and VDR siRNA transfected cells. 5,000 cells after transfection were plated on 24 well ultra-low attachment plates and total numbers of mammospheres formed were analyzed (**, p≤0.01). D, Over-expression of full length human VDR gene and analysis of VDR, Snail and E-cad protein expression by western blot. E, Analysis of EMT-specific gene signature in cells transfected either with control vector or full-length human VDR (*, p≤0.05; **, p≤0.01). F, Analysis of mammospheres forming capability in cells transfected either with control vector or VDR over-expressing (VDR OE) plasmid. 5,000 cells after transfection were plated on 24 well ultra-low attachment plates and total numbers of mammospheres formed were analyzed (**, p≤0.01).

    Techniques Used: Inhibition, Expressing, Transfection, Western Blot, Over Expression, Plasmid Preparation

    A : Left Panel: Bright field photomicrograph of mammospheres isolated from SKBR3 CELLS. Right Panel: Fluorescent micrographs of SKBR3 mammospheres stained with CD44, ESA, and CD24 antigens (green) and DAPI (blue). B, Real-time quantitative PCR analysis of c-Myc, Klf4, Oct4, and Sox2 in SKBR3 cells grown under high attachment, plastic (plas) or mammosphere (mam) conditions (*, p≤0.05; **, p≤0.01). C , Isolation of ALDH + and ALDH − population from SKBR3 cells and analysis of their mammosphere forming capacity. D, Analysis of tumor volume in nude mice by SKBR3 mammospheres and plastic cells. 1×10 5 cells obtained from dissociated mam or plas were injected into the humanized mammary fat pads of female nude mice and tumor volumes were analyzed at various time points (1–8 weeks).
    Figure Legend Snippet: A : Left Panel: Bright field photomicrograph of mammospheres isolated from SKBR3 CELLS. Right Panel: Fluorescent micrographs of SKBR3 mammospheres stained with CD44, ESA, and CD24 antigens (green) and DAPI (blue). B, Real-time quantitative PCR analysis of c-Myc, Klf4, Oct4, and Sox2 in SKBR3 cells grown under high attachment, plastic (plas) or mammosphere (mam) conditions (*, p≤0.05; **, p≤0.01). C , Isolation of ALDH + and ALDH − population from SKBR3 cells and analysis of their mammosphere forming capacity. D, Analysis of tumor volume in nude mice by SKBR3 mammospheres and plastic cells. 1×10 5 cells obtained from dissociated mam or plas were injected into the humanized mammary fat pads of female nude mice and tumor volumes were analyzed at various time points (1–8 weeks).

    Techniques Used: Isolation, Staining, Real-time Polymerase Chain Reaction, Mouse Assay, Injection

    A, Effect of 1,25D treatment on MCF7 (left) and SKBR3 (right) cell proliferation. Cells were treated with 1,25D (0–0.1 nM) and allowed to proliferate for 4 days under high attachment conditions and cell numbers were counted by trypan blue method (*, p≤0.05; **, p≤0.01). Medium was replaced after every 48 hrs with appropriate concentrations of 1,25D. B, SKBR3 (2×10 4 ) cells were plated under mammosphere conditions on a 12-well ultra-low attachment plates in presence of different concentrations of 1,25D (0–0.1 nM) and allowed to grow under mammosphere conditions for 4 or 7 days and sphere diameters were measured. Appropriate concentrations of 1,25D were additionally supplemented in the culture medium after every 48 hours. Left Panel: Micrographs were taken at 100×magnification. Right Panel: Quantitative analysis of average diameter computed from 20 different fields from each treatment group. C, Quantitative real-time PCR analysis of Cyp27 B1 and Cyp24A1 mRNA expression from MCF-7 cells grown under plas or mam conditions after 4 days of plating (**, p≤0.01). D, HPLC analysis of 24,25D3 and 1,25D synthesis in cells grown under plastic or mammos conditions from MCF-7 cells after 4 days of plating (*, p≤0.05; **, p≤0.01).
    Figure Legend Snippet: A, Effect of 1,25D treatment on MCF7 (left) and SKBR3 (right) cell proliferation. Cells were treated with 1,25D (0–0.1 nM) and allowed to proliferate for 4 days under high attachment conditions and cell numbers were counted by trypan blue method (*, p≤0.05; **, p≤0.01). Medium was replaced after every 48 hrs with appropriate concentrations of 1,25D. B, SKBR3 (2×10 4 ) cells were plated under mammosphere conditions on a 12-well ultra-low attachment plates in presence of different concentrations of 1,25D (0–0.1 nM) and allowed to grow under mammosphere conditions for 4 or 7 days and sphere diameters were measured. Appropriate concentrations of 1,25D were additionally supplemented in the culture medium after every 48 hours. Left Panel: Micrographs were taken at 100×magnification. Right Panel: Quantitative analysis of average diameter computed from 20 different fields from each treatment group. C, Quantitative real-time PCR analysis of Cyp27 B1 and Cyp24A1 mRNA expression from MCF-7 cells grown under plas or mam conditions after 4 days of plating (**, p≤0.01). D, HPLC analysis of 24,25D3 and 1,25D synthesis in cells grown under plastic or mammos conditions from MCF-7 cells after 4 days of plating (*, p≤0.05; **, p≤0.01).

    Techniques Used: Real-time Polymerase Chain Reaction, Expressing, High Performance Liquid Chromatography

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    ATCC skbr3
    PUVA therapy inhibits ErbB2 signaling. BT474, <t>SKBR3</t> and MCF7 cells were subjected to the indicated treatment conditions as described in Figure 1 . Western blot analysis was performed on whole cell lysates. Actin steady-state protein levels served as a control to ensure for equal loading of protein. Results are representative of three independent experiments.
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    PUVA therapy inhibits ErbB2 signaling. BT474, SKBR3 and MCF7 cells were subjected to the indicated treatment conditions as described in Figure 1 . Western blot analysis was performed on whole cell lysates. Actin steady-state protein levels served as a control to ensure for equal loading of protein. Results are representative of three independent experiments.

    Journal: PLoS ONE

    Article Title: Photo-Activated Psoralen Binds the ErbB2 Catalytic Kinase Domain, Blocking ErbB2 Signaling and Triggering Tumor Cell Apoptosis

    doi: 10.1371/journal.pone.0088983

    Figure Lengend Snippet: PUVA therapy inhibits ErbB2 signaling. BT474, SKBR3 and MCF7 cells were subjected to the indicated treatment conditions as described in Figure 1 . Western blot analysis was performed on whole cell lysates. Actin steady-state protein levels served as a control to ensure for equal loading of protein. Results are representative of three independent experiments.

    Article Snippet: Cell Culture and Reagents ErbB2+ (BT474; SKBR3) and ErbB2 negative (MCF-7; T47D) human breast cancer cell lines, and the human foreskin fibroblast (HFF) cell line were obtained from the American Type Culture Collection (Manassas, VA).

    Techniques: Western Blot

    Inhibition of ErbB2 signaling in response to PUVA is independent of DNA crosslinking. The chemical structures of (A) 8MOP, and (B) 7-methylpyridopsoralen (SMSF032310), which is a derivative of 8MOP that lacks the ability to crosslink DNA, are shown. (C) BT474 and SKBR3 cells were exposed to the indicated treatments. Cell growth and viability assays were performed after 72 hr. P

    Journal: PLoS ONE

    Article Title: Photo-Activated Psoralen Binds the ErbB2 Catalytic Kinase Domain, Blocking ErbB2 Signaling and Triggering Tumor Cell Apoptosis

    doi: 10.1371/journal.pone.0088983

    Figure Lengend Snippet: Inhibition of ErbB2 signaling in response to PUVA is independent of DNA crosslinking. The chemical structures of (A) 8MOP, and (B) 7-methylpyridopsoralen (SMSF032310), which is a derivative of 8MOP that lacks the ability to crosslink DNA, are shown. (C) BT474 and SKBR3 cells were exposed to the indicated treatments. Cell growth and viability assays were performed after 72 hr. P

    Article Snippet: Cell Culture and Reagents ErbB2+ (BT474; SKBR3) and ErbB2 negative (MCF-7; T47D) human breast cancer cell lines, and the human foreskin fibroblast (HFF) cell line were obtained from the American Type Culture Collection (Manassas, VA).

    Techniques: Inhibition

    PUVA antitumor activity in HER2 + breast cancer cells. Cells were pre-treated with the indicated concentrations of 8MOP for 4 hr before UVA irradiation (2J), and then cultured for an additional 72 hr before being analyzed for cell growth (A) BT474; (B) SKBR3; (C) MCF7, (D) HFF and apoptosis (D) BT474; (E) SKBR3. Cells treated with vehicle (0.01% DMSO) alone served as controls. Results represent the mean +/− standard error of triplicate samples, and are representative of three independent experiments.

    Journal: PLoS ONE

    Article Title: Photo-Activated Psoralen Binds the ErbB2 Catalytic Kinase Domain, Blocking ErbB2 Signaling and Triggering Tumor Cell Apoptosis

    doi: 10.1371/journal.pone.0088983

    Figure Lengend Snippet: PUVA antitumor activity in HER2 + breast cancer cells. Cells were pre-treated with the indicated concentrations of 8MOP for 4 hr before UVA irradiation (2J), and then cultured for an additional 72 hr before being analyzed for cell growth (A) BT474; (B) SKBR3; (C) MCF7, (D) HFF and apoptosis (D) BT474; (E) SKBR3. Cells treated with vehicle (0.01% DMSO) alone served as controls. Results represent the mean +/− standard error of triplicate samples, and are representative of three independent experiments.

    Article Snippet: Cell Culture and Reagents ErbB2+ (BT474; SKBR3) and ErbB2 negative (MCF-7; T47D) human breast cancer cell lines, and the human foreskin fibroblast (HFF) cell line were obtained from the American Type Culture Collection (Manassas, VA).

    Techniques: Activity Assay, Irradiation, Cell Culture

    The combination of PUVA with the irreversible pan-ErbB TKI neratinib results in enhanced antitumor activity. The growth and viability of BT474 and SKBR3 cells (top bar graphs) after being subjected to the indicated treatment conditions. The combination of PUVA plus neratinib: P

    Journal: PLoS ONE

    Article Title: Photo-Activated Psoralen Binds the ErbB2 Catalytic Kinase Domain, Blocking ErbB2 Signaling and Triggering Tumor Cell Apoptosis

    doi: 10.1371/journal.pone.0088983

    Figure Lengend Snippet: The combination of PUVA with the irreversible pan-ErbB TKI neratinib results in enhanced antitumor activity. The growth and viability of BT474 and SKBR3 cells (top bar graphs) after being subjected to the indicated treatment conditions. The combination of PUVA plus neratinib: P

    Article Snippet: Cell Culture and Reagents ErbB2+ (BT474; SKBR3) and ErbB2 negative (MCF-7; T47D) human breast cancer cell lines, and the human foreskin fibroblast (HFF) cell line were obtained from the American Type Culture Collection (Manassas, VA).

    Techniques: Activity Assay

    pTyr397FAK expression is higher in triple negative cell lines and ALDH + cells. FAK inhibition in combination with adjuvant therapies reduces CSC activity. a Representative Western blot demonstrating baseline expression of pTyr397FAK, FAK, and GAPDH in breast cell lines including invasive carcinoma cell lines reflecting all molecular phenotypes. b Illustrative plot of relative density of pTyr397FAK to FAK, with relative density of pTyr397FAK to FAK measured and corrected for GAPDH. pTyr397FAK expression in normal ductal cell line MCF10a used as comparison. One-way ANOVA with post hoc Dunnett’s test ( n = 2 for MCF10a and DCIS.com and n = 3 for IDC cell lines). c Representative Western blot demonstrating pTyr397FAK expression in ALDH + and ALDH − MDA-MB-231 cells with illustrative plot of relative densities shown in ( d ) ( n = 3). e ALDH + cells have increased primary MFE compared to ALDH − expressing cells. Both D E analysed using unpaired two tailed t -test. f FAK inhibition with VS4718 0.5 μM reduced primary MFE as a single agent therapy across all cell lines. This was evaluated alongside 1 μM of Tamoxifen in MCF7, 0.1 μM of Lapatinib in BT474 and SKBr3 cells and 0.1 μM Paclitaxel in MDA-MB-231 and SUM159 cells. Each experiment had a minimum of five biological repeats and six technical replicates. g FAK inhibition reduced mammosphere self-renewal in SKBr3 and SUM159 cells when used as monotherapy and in MDA-MB-231 cells when combined with Paclitaxel. All error bars are mean + SEM. Two-way ANOVA with post hoc Tukey’s test (ns not significant, * p

    Journal: NPJ Breast Cancer

    Article Title: FAK inhibition alone or in combination with adjuvant therapies reduces cancer stem cell activity

    doi: 10.1038/s41523-021-00263-3

    Figure Lengend Snippet: pTyr397FAK expression is higher in triple negative cell lines and ALDH + cells. FAK inhibition in combination with adjuvant therapies reduces CSC activity. a Representative Western blot demonstrating baseline expression of pTyr397FAK, FAK, and GAPDH in breast cell lines including invasive carcinoma cell lines reflecting all molecular phenotypes. b Illustrative plot of relative density of pTyr397FAK to FAK, with relative density of pTyr397FAK to FAK measured and corrected for GAPDH. pTyr397FAK expression in normal ductal cell line MCF10a used as comparison. One-way ANOVA with post hoc Dunnett’s test ( n = 2 for MCF10a and DCIS.com and n = 3 for IDC cell lines). c Representative Western blot demonstrating pTyr397FAK expression in ALDH + and ALDH − MDA-MB-231 cells with illustrative plot of relative densities shown in ( d ) ( n = 3). e ALDH + cells have increased primary MFE compared to ALDH − expressing cells. Both D E analysed using unpaired two tailed t -test. f FAK inhibition with VS4718 0.5 μM reduced primary MFE as a single agent therapy across all cell lines. This was evaluated alongside 1 μM of Tamoxifen in MCF7, 0.1 μM of Lapatinib in BT474 and SKBr3 cells and 0.1 μM Paclitaxel in MDA-MB-231 and SUM159 cells. Each experiment had a minimum of five biological repeats and six technical replicates. g FAK inhibition reduced mammosphere self-renewal in SKBr3 and SUM159 cells when used as monotherapy and in MDA-MB-231 cells when combined with Paclitaxel. All error bars are mean + SEM. Two-way ANOVA with post hoc Tukey’s test (ns not significant, * p

    Article Snippet: MCF7, BT474, SKBr3, MDA-MB231, and SUM159 lines were purchased from ATCC.

    Techniques: Expressing, Inhibition, Activity Assay, Western Blot, Multiple Displacement Amplification, Two Tailed Test

    Effects of FAE on mitochondrial membrane potential and Caspase activation in breast cancer cells and normal cells. (A) The breast cancer cells, MDAMB231, T47D, SKBr3 and normal cells like Human Mammary Epithelial cells and Human Endothelial cells after treatment with FAE at 100 µg/ml for 24 h were stained with 50 nM of TMRM and 0.5 µg/ml of Hoechst 33342 for 15 mins. Then the cells were imaged under fluorescent microscope using DAPI and Rhodamine filter sets using 40× objective. The images were captured with Retiga Exi camera using NIS element software (Nikon). (B) Ac-LEHD-AFC cleavage (Caspase 9 activity) (C) Ac-DEVD-AMC cleavage (Caspase 3/7 activity). MCF-7 cells were treated at IC 50 value of FAE for 24, 36 and 48 h. The results were measured fluorometrically. Values are expressed as mean ± SD of triplicate samples. Significant difference from control value was indicated by *(p

    Journal: PLoS ONE

    Article Title: Bax Translocation Mediated Mitochondrial Apoptosis and Caspase Dependent Photosensitizing Effect of Ficus religiosa on Cancer Cells

    doi: 10.1371/journal.pone.0040055

    Figure Lengend Snippet: Effects of FAE on mitochondrial membrane potential and Caspase activation in breast cancer cells and normal cells. (A) The breast cancer cells, MDAMB231, T47D, SKBr3 and normal cells like Human Mammary Epithelial cells and Human Endothelial cells after treatment with FAE at 100 µg/ml for 24 h were stained with 50 nM of TMRM and 0.5 µg/ml of Hoechst 33342 for 15 mins. Then the cells were imaged under fluorescent microscope using DAPI and Rhodamine filter sets using 40× objective. The images were captured with Retiga Exi camera using NIS element software (Nikon). (B) Ac-LEHD-AFC cleavage (Caspase 9 activity) (C) Ac-DEVD-AMC cleavage (Caspase 3/7 activity). MCF-7 cells were treated at IC 50 value of FAE for 24, 36 and 48 h. The results were measured fluorometrically. Values are expressed as mean ± SD of triplicate samples. Significant difference from control value was indicated by *(p

    Article Snippet: Cell Culture The breast cancer cell lines MCF-7, T47D, SKBr3 and MDAMB 231 and the normal breast epithelial cell, MCF-10A were obtained from the ATCC (American Type Culture collection, Manassas, VA).

    Techniques: Activation Assay, Staining, Microscopy, Software, Activity Assay

    Effects of CAXII inhibitors on intracellular doxorubicin retention in drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicinsensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were grown for 24 h in the presence of 5 μM doxorubicin, alone (–) or in the presence of 5 nM of compounds 1–8. Tariquidar (25 nM; Tar) was included as Pgp inhibitor. The intracellular drug content was measured fluorimetrically. Data are presented as means ± SD ( n = 4). Versus doxorubicin alone (–): *p

    Journal: Oncotarget

    Article Title: P-glycoprotein-mediated chemoresistance is reversed by carbonic anhydrase XII inhibitors

    doi: 10.18632/oncotarget.13040

    Figure Lengend Snippet: Effects of CAXII inhibitors on intracellular doxorubicin retention in drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicinsensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were grown for 24 h in the presence of 5 μM doxorubicin, alone (–) or in the presence of 5 nM of compounds 1–8. Tariquidar (25 nM; Tar) was included as Pgp inhibitor. The intracellular drug content was measured fluorimetrically. Data are presented as means ± SD ( n = 4). Versus doxorubicin alone (–): *p

    Article Snippet: Cell linesHuman colon cancer HT29 cells, lung cancer A549 cells, breast cancer MCF-7, SKBR3, T74D and MDA-MB-231 cells, osteosarcoma U2OS and SaOS cells, murine chemoresistant JC cells, not-transformed human colon epithelial CCD-Co-18 cells, lung epithelial BEAS-2B cells, breast epithelial MCF10A cells were purchased from ATCC (Manassas, VA).

    Techniques: Multiple Displacement Amplification

    Effects of CAXII inhibitors on viability of drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicin-sensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were incubated for 72 h with increasing concentrations (1 nM – 1 mM) of doxorubicin (dox), alone or in the presence of 5 nM of compound 1, 2 and 4, then stained in quadruplicate with neutral red. Tariquidar (25 nM; Tar) was included as a Pgp inhibitor. IC 50 of tariquidar alone was > 10 μM; at 25 nM tariquidar reduced viability ≤ 8.23 ± 1.44 % in each cell line. Data are presented as mean IC 50 ± SD ( n = 4). HT29/DX, A549/DX, U2OS/DX, SaOS/DX versus HT29, A549, U2OS, SaOS cells: * p

    Journal: Oncotarget

    Article Title: P-glycoprotein-mediated chemoresistance is reversed by carbonic anhydrase XII inhibitors

    doi: 10.18632/oncotarget.13040

    Figure Lengend Snippet: Effects of CAXII inhibitors on viability of drug-sensitive and drug-resistant cancer cells Human doxorubicin-sensitive colon cancer HT29 cells and their resistant counterpart HT29/DX cells (panel A ), human doxorubicin-sensitive lung cancer A549 cells and their resistant counterpart A549/DX cells (panel B ), human doxorubicin-sensitive and resistant breast cancer MCF7 (panel C ), SKBR3 (panel D ), T74D (panel E ) and MDA-MB-231 cells (panel F ), murine doxorubicin-resistant TUBO (panel G ) and JC cells (panel H ), human doxorubicin-sensitive osteosarcoma U2OS cells and their resistant counterpart U2OS/DX cells (panel I ), human doxorubicin-sensitive osteosarcoma Saos and their resistant counterpart SaOS/DX (panel J ) were incubated for 72 h with increasing concentrations (1 nM – 1 mM) of doxorubicin (dox), alone or in the presence of 5 nM of compound 1, 2 and 4, then stained in quadruplicate with neutral red. Tariquidar (25 nM; Tar) was included as a Pgp inhibitor. IC 50 of tariquidar alone was > 10 μM; at 25 nM tariquidar reduced viability ≤ 8.23 ± 1.44 % in each cell line. Data are presented as mean IC 50 ± SD ( n = 4). HT29/DX, A549/DX, U2OS/DX, SaOS/DX versus HT29, A549, U2OS, SaOS cells: * p

    Article Snippet: Cell linesHuman colon cancer HT29 cells, lung cancer A549 cells, breast cancer MCF-7, SKBR3, T74D and MDA-MB-231 cells, osteosarcoma U2OS and SaOS cells, murine chemoresistant JC cells, not-transformed human colon epithelial CCD-Co-18 cells, lung epithelial BEAS-2B cells, breast epithelial MCF10A cells were purchased from ATCC (Manassas, VA).

    Techniques: Multiple Displacement Amplification, Incubation, Staining