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methylophilus methylotrophus atcc 53528  (ATCC)


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    ATCC methylophilus methylotrophus atcc 53528
    Methylophilus Methylotrophus Atcc 53528, supplied by ATCC, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 92 stars, based on 1 article reviews
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    Cell Signaling Technology Inc antibodies against foxa1 hnf3α
    Effect of telaprevir on the proliferation of nontransformed and transformed cell lines. (A) Telaprevir (Tel) inhibitor concentration 50 (IC50‐μ m ) measured on a panel of nontransformed (i.e., human fibroblast and MCF10a cells) and transformed (i.e., MDA‐MB‐231, MCF‐7, T47D‐1, BT‐474, SKBR3, AU565, HeLa, U251, SKOV3, and DU145 cells) cell lines at 5 days. N = 4. (B) The panel indicates cell sensitivity to Tel antiproliferative effect (red) and ERα (blue), HER2 (orange), and <t>FOXA1</t> (green) mRNA expression in the indicated cell lines. ERα, Her2, and FOXA1 expression was extrapolated in silico from the Depmap portal database ( https://depmap.org/portal/ ). (C) Real‐time growth curves in the indicated cell lines treated with Tel 20 μ m (0–96 h). Normalized cell index (i.e., CI) has been calculated at each time point with respect to the control sample (gray line). Data are the mean ± standard errors n = 8. Real‐time growth curve analyses in MCF‐7 (D), SKBR3 (E), and SKOV3 (F) cells depleted of ERα (red line) and/or FOXA1 (green line) by siRNA. Normalized cell index (i.e., CI) has been calculated at each time point with respect to the control sample (gray line). Data are the mean ± standard errors n = 8. (G) Tel inhibition concentration 50 (IC 50 ‐μ m ) on SKBR3 and SKOV3 cells proliferation measured in the presence or the absence of FOXA1 expression at 3 days in SKBR3 cells and 5 days in SKOV3 cells. Data are the mean ± standard deviations with a P ‐value < 0.0001. **** indicates significant differences with respect to the control sample (siRNA CTR or CTR) calculated with Student t ‐test. n = 17. [Colour figure can be viewed at wileyonlinelibrary.com ]
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    Cell Signaling Technology Inc foxa1
    A GSEA showed that genes induced by <t>FOXA1</t> were inhibited by ivermectin in C4-2 cells. B RT-qPCR analysis of FOXA1 induced genes ( CDKN3 , CDCA2 , and CAMKK2 ) and FOXA1 repressed EMT-associated genes ( MET , MMP7 , and SOX9 ) in C4-2 cells treated with ivermectin for 48 h. C Western blot analysis of FOXA1 and N-cadherin in LNCaP and C4-2 cells treated with ivermectin for 48 h. D ChIP–qPCR analysis for FOXA1 or AR occupancy, and FAIRE–qPCR analysis of chromatin accessibility at a target regulated by AR and FOXA1 (KLK3 and NKX3-1) in C4-2 cells treated with ivermectin. E ChIP–qPCR analysis for FOXA1 and FAIRE-PCR analysis of chromatin accessibility at a target regulated by FOXA1 (E2F1 and MET) in C4-2 cells treated with ivermectin. F FOXA1 knockdown impaired the ivermectin-repressed expression of KLK3 and E2F1 genes. mRNA levels were measured 48 h after the implementation of the ivermectin treatment and siRNA transfection by RT-qPCR in C4-2 cells. G , H Western blots showing thermostable FOXA1 and AR following indicated heat shocks in the presence (+) or absence (−) of 50 μM ivermectin in LNCaP ( G ) and C4-2 ( H ) cells. I Western blots showing thermostable FOXA1 following indicated heat shocks in the presence (+) or absence (−) of 50 μM ivermectin in 22RV1 cells. J GSEA showed the inactivation of FOXA1 induced genes in 22RV1 cells after the ivermectin treatment. K RT-qPCR analysis of FL-AR and ARv7 in 22RV1 cells treated with ivermectin for 48 h. L ChIP–qPCR analysis for FOXA1 and FAIRE–qPCR analysis of chromatin accessibility at KLK3 and E2F1 in 22RV1 cells treated with ivermectin.
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    Effect of telaprevir on the proliferation of nontransformed and transformed cell lines. (A) Telaprevir (Tel) inhibitor concentration 50 (IC50‐μ m ) measured on a panel of nontransformed (i.e., human fibroblast and MCF10a cells) and transformed (i.e., MDA‐MB‐231, MCF‐7, T47D‐1, BT‐474, SKBR3, AU565, HeLa, U251, SKOV3, and DU145 cells) cell lines at 5 days. N = 4. (B) The panel indicates cell sensitivity to Tel antiproliferative effect (red) and ERα (blue), HER2 (orange), and FOXA1 (green) mRNA expression in the indicated cell lines. ERα, Her2, and FOXA1 expression was extrapolated in silico from the Depmap portal database ( https://depmap.org/portal/ ). (C) Real‐time growth curves in the indicated cell lines treated with Tel 20 μ m (0–96 h). Normalized cell index (i.e., CI) has been calculated at each time point with respect to the control sample (gray line). Data are the mean ± standard errors n = 8. Real‐time growth curve analyses in MCF‐7 (D), SKBR3 (E), and SKOV3 (F) cells depleted of ERα (red line) and/or FOXA1 (green line) by siRNA. Normalized cell index (i.e., CI) has been calculated at each time point with respect to the control sample (gray line). Data are the mean ± standard errors n = 8. (G) Tel inhibition concentration 50 (IC 50 ‐μ m ) on SKBR3 and SKOV3 cells proliferation measured in the presence or the absence of FOXA1 expression at 3 days in SKBR3 cells and 5 days in SKOV3 cells. Data are the mean ± standard deviations with a P ‐value < 0.0001. **** indicates significant differences with respect to the control sample (siRNA CTR or CTR) calculated with Student t ‐test. n = 17. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Journal: Molecular Oncology

    Article Title: The antiviral drug telaprevir induces cell death by reducing FOXA1 expression in estrogen receptor α ( ERα )‐positive breast cancer cells

    doi: 10.1002/1878-0261.13303

    Figure Lengend Snippet: Effect of telaprevir on the proliferation of nontransformed and transformed cell lines. (A) Telaprevir (Tel) inhibitor concentration 50 (IC50‐μ m ) measured on a panel of nontransformed (i.e., human fibroblast and MCF10a cells) and transformed (i.e., MDA‐MB‐231, MCF‐7, T47D‐1, BT‐474, SKBR3, AU565, HeLa, U251, SKOV3, and DU145 cells) cell lines at 5 days. N = 4. (B) The panel indicates cell sensitivity to Tel antiproliferative effect (red) and ERα (blue), HER2 (orange), and FOXA1 (green) mRNA expression in the indicated cell lines. ERα, Her2, and FOXA1 expression was extrapolated in silico from the Depmap portal database ( https://depmap.org/portal/ ). (C) Real‐time growth curves in the indicated cell lines treated with Tel 20 μ m (0–96 h). Normalized cell index (i.e., CI) has been calculated at each time point with respect to the control sample (gray line). Data are the mean ± standard errors n = 8. Real‐time growth curve analyses in MCF‐7 (D), SKBR3 (E), and SKOV3 (F) cells depleted of ERα (red line) and/or FOXA1 (green line) by siRNA. Normalized cell index (i.e., CI) has been calculated at each time point with respect to the control sample (gray line). Data are the mean ± standard errors n = 8. (G) Tel inhibition concentration 50 (IC 50 ‐μ m ) on SKBR3 and SKOV3 cells proliferation measured in the presence or the absence of FOXA1 expression at 3 days in SKBR3 cells and 5 days in SKOV3 cells. Data are the mean ± standard deviations with a P ‐value < 0.0001. **** indicates significant differences with respect to the control sample (siRNA CTR or CTR) calculated with Student t ‐test. n = 17. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Article Snippet: Antibodies against FOXA1/HNF3α (53528S, rabbit), phospho‐AKT Ser473 (4058S, rabbit), PARP (9542T, rabbit), phospho‐IGF1‐R (3024S, rabbit), and IGF1‐R (3027S, rabbit) were purchased by Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Transformation Assay, Concentration Assay, Expressing, In Silico, Inhibition

    FOXA1 involvement in telaprevir effect on ERα levels and functions. (A) Western blotting and relative quantitation (heat map) of FOXA1 levels in MCF‐7, BT‐474, SKBR3, and AU565 cells treated with telaprevir (Tel 20 μ m ) for 72 h. The loading control was done by evaluating vinculin expression in the same filter. (B) Western blotting and relative quantitation (heat map) of ERα and vinculin levels in MCF‐7, BT‐474, and SKOV3 cells treated with Tel 20 μ m for 72 h. Panels show representative blots. Histograms are reported in Fig. A,B, where the mean ± standard deviation with a P ‐value < 0.001 (***) and 0.0001 (****) is reported. * indicates significant differences with respect to the control sample (−) of each cell line and has been calculated with Student t ‐test. n = 4 for all the experiments while n = 6 for SKBR3 cell lines. The activity of FOXA1‐specific enhancer in ESR1 promoter region measured in MCF‐7 ESR1‐NLuc cells treated with the indicated concentrations of Tel for 48 h (C) or with the FOXA1 siRNA (E). Data are the mean ± standard deviation with a P ‐value < 0.0001 (****). * indicates significant differences with respect to the control sample (0) calculated with one‐way ANOVA followed by the Tukey post‐test (panel C) and with Student t ‐test (panel E), n = 10. (D) ERE promoter activity in MCF‐7 ERE‐NLuc cells treated with the indicated doses of Tel for 48 h both in the absence or presence of FOXA1 siRNA. n = 10. Data are the mean ± standard deviation with a P ‐value < 0.001 (***). * indicates significant differences with respect to the control sample (CTR) calculated with Student t ‐test. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Journal: Molecular Oncology

    Article Title: The antiviral drug telaprevir induces cell death by reducing FOXA1 expression in estrogen receptor α ( ERα )‐positive breast cancer cells

    doi: 10.1002/1878-0261.13303

    Figure Lengend Snippet: FOXA1 involvement in telaprevir effect on ERα levels and functions. (A) Western blotting and relative quantitation (heat map) of FOXA1 levels in MCF‐7, BT‐474, SKBR3, and AU565 cells treated with telaprevir (Tel 20 μ m ) for 72 h. The loading control was done by evaluating vinculin expression in the same filter. (B) Western blotting and relative quantitation (heat map) of ERα and vinculin levels in MCF‐7, BT‐474, and SKOV3 cells treated with Tel 20 μ m for 72 h. Panels show representative blots. Histograms are reported in Fig. A,B, where the mean ± standard deviation with a P ‐value < 0.001 (***) and 0.0001 (****) is reported. * indicates significant differences with respect to the control sample (−) of each cell line and has been calculated with Student t ‐test. n = 4 for all the experiments while n = 6 for SKBR3 cell lines. The activity of FOXA1‐specific enhancer in ESR1 promoter region measured in MCF‐7 ESR1‐NLuc cells treated with the indicated concentrations of Tel for 48 h (C) or with the FOXA1 siRNA (E). Data are the mean ± standard deviation with a P ‐value < 0.0001 (****). * indicates significant differences with respect to the control sample (0) calculated with one‐way ANOVA followed by the Tukey post‐test (panel C) and with Student t ‐test (panel E), n = 10. (D) ERE promoter activity in MCF‐7 ERE‐NLuc cells treated with the indicated doses of Tel for 48 h both in the absence or presence of FOXA1 siRNA. n = 10. Data are the mean ± standard deviation with a P ‐value < 0.001 (***). * indicates significant differences with respect to the control sample (CTR) calculated with Student t ‐test. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Article Snippet: Antibodies against FOXA1/HNF3α (53528S, rabbit), phospho‐AKT Ser473 (4058S, rabbit), PARP (9542T, rabbit), phospho‐IGF1‐R (3024S, rabbit), and IGF1‐R (3027S, rabbit) were purchased by Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Western Blot, Quantitation Assay, Expressing, Standard Deviation, Activity Assay

    IGF1‐R/AKT/FOXA1 involvement in telaprevir mechanism of action in MCF‐7 cells. (A) Western blotting analyses of pAKT, AKT, IGF1‐R, FOXA1, ERα and vinculin protein levels in MCF‐7 cells treated with the indicated doses of Tel for 48 h. Densitometric analysis is shown in Fig. C. n = 3 for pAKT and IGF‐1R, n = 4 for FOXA1, n = 5 for ERα. (B) Western blotting analyses of pAKT, AKT and vinculin protein levels in MCF‐7 cells treated with telaprevir (Tel ‐ 20 μ m ) at the indicated time points. Densitometric analysis and number of replicates are shown in Fig. D. n = 3 for 0.25 and 0.5 h, n = 4 for 1 and 3 h, n = 11 for 6 h, n = 6 for 24 and 48 h. (C) Western blotting analyses of pAKT, AKT and vinculin protein levels in MCF‐7 cells pretreated with NVP AEW541 (NVP 1 μ m ) for 6 h and then treated with Tel (20 μ m ‐6 h). Densitometric analysis and number of replicates are shown in Fig. E. n = 6 (D, E, D’, E’) Western blotting analyses and relative densitometric analyses of IGF1‐R, ERα and vinculin protein levels in MCF‐7 cell lysates incubated with increasing doses of telaprevir (Tel 1–100 μ m ) and NVP AEW541 (NVP 1–100 μ m ) in the presence or absence of pronase (1 : 1000). Data are the mean ± standard deviation n = 3. ** ( P ‐value < 0.01) or * ( P ‐value < 0.05) indicates significant differences with respect to the control sample (0) calculated with one‐way ANOVA followed by the Tukey post‐test. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Journal: Molecular Oncology

    Article Title: The antiviral drug telaprevir induces cell death by reducing FOXA1 expression in estrogen receptor α ( ERα )‐positive breast cancer cells

    doi: 10.1002/1878-0261.13303

    Figure Lengend Snippet: IGF1‐R/AKT/FOXA1 involvement in telaprevir mechanism of action in MCF‐7 cells. (A) Western blotting analyses of pAKT, AKT, IGF1‐R, FOXA1, ERα and vinculin protein levels in MCF‐7 cells treated with the indicated doses of Tel for 48 h. Densitometric analysis is shown in Fig. C. n = 3 for pAKT and IGF‐1R, n = 4 for FOXA1, n = 5 for ERα. (B) Western blotting analyses of pAKT, AKT and vinculin protein levels in MCF‐7 cells treated with telaprevir (Tel ‐ 20 μ m ) at the indicated time points. Densitometric analysis and number of replicates are shown in Fig. D. n = 3 for 0.25 and 0.5 h, n = 4 for 1 and 3 h, n = 11 for 6 h, n = 6 for 24 and 48 h. (C) Western blotting analyses of pAKT, AKT and vinculin protein levels in MCF‐7 cells pretreated with NVP AEW541 (NVP 1 μ m ) for 6 h and then treated with Tel (20 μ m ‐6 h). Densitometric analysis and number of replicates are shown in Fig. E. n = 6 (D, E, D’, E’) Western blotting analyses and relative densitometric analyses of IGF1‐R, ERα and vinculin protein levels in MCF‐7 cell lysates incubated with increasing doses of telaprevir (Tel 1–100 μ m ) and NVP AEW541 (NVP 1–100 μ m ) in the presence or absence of pronase (1 : 1000). Data are the mean ± standard deviation n = 3. ** ( P ‐value < 0.01) or * ( P ‐value < 0.05) indicates significant differences with respect to the control sample (0) calculated with one‐way ANOVA followed by the Tukey post‐test. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Article Snippet: Antibodies against FOXA1/HNF3α (53528S, rabbit), phospho‐AKT Ser473 (4058S, rabbit), PARP (9542T, rabbit), phospho‐IGF1‐R (3024S, rabbit), and IGF1‐R (3027S, rabbit) were purchased by Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Western Blot, Incubation, Standard Deviation

    Analysis of telaprevir‐induced apoptosis. (A) Western blotting and (A’) relative densitometric analyses of cleaved PARP and tubulin (Tub) in MCF‐7 cells treated with different doses of telaprevir (Tel 10–40 μ m ) for 48 h and staurosporine (STS 100 n m ) for 24 h. Data are the mean ± standard deviation with P ‐value < 0.0001. n = 4 for all samples but for 40 μ m where n = 3. **** indicates significant differences with respect to the control sample (0) calculated with one‐way ANOVA followed by the Tukey post‐test. Western blotting analyses of telaprevir (Tel) effect on cleaved PARP, FOXA1 and tubulin (Tub) levels in the presence of FOXA1 siRNA (B) and NVP (C). Cells were transfected with FOXA1 siRNA and treated with 1 μ m NVP before 48 h of 20 μ m Tel treatment. (D) Western blotting analyses of cleaved PARP, FOXA1 and vinculin expression in MCF‐7 cells pretreated with different doses of IGF (10–250 ng·mL −1 ) for 1 h before 48 h of 20 μ m Tel treatment. Densitometric analyses of panel (B) (C) and (D) are shown in Fig. C–H. n = 4 for (B) (C) and (D), n = 3 for FOXA1 levels in panel D. (E–H) Western blotting analyses of cleaved PARP and tubulin in HeLa, SKOV3, MCF‐7, and SKBR3 cells treated with telaprevir (Tel 20 μ m ) for 48 h. Densitometric analysis are shown in Fig. B. n = 3 for all the cell lines and n = 4 for SKOV3 cells.

    Journal: Molecular Oncology

    Article Title: The antiviral drug telaprevir induces cell death by reducing FOXA1 expression in estrogen receptor α ( ERα )‐positive breast cancer cells

    doi: 10.1002/1878-0261.13303

    Figure Lengend Snippet: Analysis of telaprevir‐induced apoptosis. (A) Western blotting and (A’) relative densitometric analyses of cleaved PARP and tubulin (Tub) in MCF‐7 cells treated with different doses of telaprevir (Tel 10–40 μ m ) for 48 h and staurosporine (STS 100 n m ) for 24 h. Data are the mean ± standard deviation with P ‐value < 0.0001. n = 4 for all samples but for 40 μ m where n = 3. **** indicates significant differences with respect to the control sample (0) calculated with one‐way ANOVA followed by the Tukey post‐test. Western blotting analyses of telaprevir (Tel) effect on cleaved PARP, FOXA1 and tubulin (Tub) levels in the presence of FOXA1 siRNA (B) and NVP (C). Cells were transfected with FOXA1 siRNA and treated with 1 μ m NVP before 48 h of 20 μ m Tel treatment. (D) Western blotting analyses of cleaved PARP, FOXA1 and vinculin expression in MCF‐7 cells pretreated with different doses of IGF (10–250 ng·mL −1 ) for 1 h before 48 h of 20 μ m Tel treatment. Densitometric analyses of panel (B) (C) and (D) are shown in Fig. C–H. n = 4 for (B) (C) and (D), n = 3 for FOXA1 levels in panel D. (E–H) Western blotting analyses of cleaved PARP and tubulin in HeLa, SKOV3, MCF‐7, and SKBR3 cells treated with telaprevir (Tel 20 μ m ) for 48 h. Densitometric analysis are shown in Fig. B. n = 3 for all the cell lines and n = 4 for SKOV3 cells.

    Article Snippet: Antibodies against FOXA1/HNF3α (53528S, rabbit), phospho‐AKT Ser473 (4058S, rabbit), PARP (9542T, rabbit), phospho‐IGF1‐R (3024S, rabbit), and IGF1‐R (3027S, rabbit) were purchased by Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Western Blot, Standard Deviation, Transfection, Expressing

    Telaprevir antiproliferative effect is correlated to the ratio of FOXA1/IGF1‐R expression. (A) Bar graph of telaprevir (Tel) inhibition concentration 50 (IC 50 ‐μ m ) value measured in the indicated cancer cell lines at 5 days. Data are the means ± the SD n = 3. (B) Western blotting analyses of FOXA1, IGF1‐R and vinculin protein levels in the indicated cancer cell lines. This blot has been performed twice. (C) The scatter plot of Spearman correlation analyses for the ratio of FOXA1/IGF1‐R protein levels and Tel IC 50 (μ m ) on the proliferation of the above‐mentioned cancer cell lines. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Journal: Molecular Oncology

    Article Title: The antiviral drug telaprevir induces cell death by reducing FOXA1 expression in estrogen receptor α ( ERα )‐positive breast cancer cells

    doi: 10.1002/1878-0261.13303

    Figure Lengend Snippet: Telaprevir antiproliferative effect is correlated to the ratio of FOXA1/IGF1‐R expression. (A) Bar graph of telaprevir (Tel) inhibition concentration 50 (IC 50 ‐μ m ) value measured in the indicated cancer cell lines at 5 days. Data are the means ± the SD n = 3. (B) Western blotting analyses of FOXA1, IGF1‐R and vinculin protein levels in the indicated cancer cell lines. This blot has been performed twice. (C) The scatter plot of Spearman correlation analyses for the ratio of FOXA1/IGF1‐R protein levels and Tel IC 50 (μ m ) on the proliferation of the above‐mentioned cancer cell lines. [Colour figure can be viewed at wileyonlinelibrary.com ]

    Article Snippet: Antibodies against FOXA1/HNF3α (53528S, rabbit), phospho‐AKT Ser473 (4058S, rabbit), PARP (9542T, rabbit), phospho‐IGF1‐R (3024S, rabbit), and IGF1‐R (3027S, rabbit) were purchased by Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Expressing, Inhibition, Concentration Assay, Western Blot

    A GSEA showed that genes induced by FOXA1 were inhibited by ivermectin in C4-2 cells. B RT-qPCR analysis of FOXA1 induced genes ( CDKN3 , CDCA2 , and CAMKK2 ) and FOXA1 repressed EMT-associated genes ( MET , MMP7 , and SOX9 ) in C4-2 cells treated with ivermectin for 48 h. C Western blot analysis of FOXA1 and N-cadherin in LNCaP and C4-2 cells treated with ivermectin for 48 h. D ChIP–qPCR analysis for FOXA1 or AR occupancy, and FAIRE–qPCR analysis of chromatin accessibility at a target regulated by AR and FOXA1 (KLK3 and NKX3-1) in C4-2 cells treated with ivermectin. E ChIP–qPCR analysis for FOXA1 and FAIRE-PCR analysis of chromatin accessibility at a target regulated by FOXA1 (E2F1 and MET) in C4-2 cells treated with ivermectin. F FOXA1 knockdown impaired the ivermectin-repressed expression of KLK3 and E2F1 genes. mRNA levels were measured 48 h after the implementation of the ivermectin treatment and siRNA transfection by RT-qPCR in C4-2 cells. G , H Western blots showing thermostable FOXA1 and AR following indicated heat shocks in the presence (+) or absence (−) of 50 μM ivermectin in LNCaP ( G ) and C4-2 ( H ) cells. I Western blots showing thermostable FOXA1 following indicated heat shocks in the presence (+) or absence (−) of 50 μM ivermectin in 22RV1 cells. J GSEA showed the inactivation of FOXA1 induced genes in 22RV1 cells after the ivermectin treatment. K RT-qPCR analysis of FL-AR and ARv7 in 22RV1 cells treated with ivermectin for 48 h. L ChIP–qPCR analysis for FOXA1 and FAIRE–qPCR analysis of chromatin accessibility at KLK3 and E2F1 in 22RV1 cells treated with ivermectin.

    Journal: Cell Death & Disease

    Article Title: Integrated analysis reveals FOXA1 and Ku70/Ku80 as targets of ivermectin in prostate cancer

    doi: 10.1038/s41419-022-05182-0

    Figure Lengend Snippet: A GSEA showed that genes induced by FOXA1 were inhibited by ivermectin in C4-2 cells. B RT-qPCR analysis of FOXA1 induced genes ( CDKN3 , CDCA2 , and CAMKK2 ) and FOXA1 repressed EMT-associated genes ( MET , MMP7 , and SOX9 ) in C4-2 cells treated with ivermectin for 48 h. C Western blot analysis of FOXA1 and N-cadherin in LNCaP and C4-2 cells treated with ivermectin for 48 h. D ChIP–qPCR analysis for FOXA1 or AR occupancy, and FAIRE–qPCR analysis of chromatin accessibility at a target regulated by AR and FOXA1 (KLK3 and NKX3-1) in C4-2 cells treated with ivermectin. E ChIP–qPCR analysis for FOXA1 and FAIRE-PCR analysis of chromatin accessibility at a target regulated by FOXA1 (E2F1 and MET) in C4-2 cells treated with ivermectin. F FOXA1 knockdown impaired the ivermectin-repressed expression of KLK3 and E2F1 genes. mRNA levels were measured 48 h after the implementation of the ivermectin treatment and siRNA transfection by RT-qPCR in C4-2 cells. G , H Western blots showing thermostable FOXA1 and AR following indicated heat shocks in the presence (+) or absence (−) of 50 μM ivermectin in LNCaP ( G ) and C4-2 ( H ) cells. I Western blots showing thermostable FOXA1 following indicated heat shocks in the presence (+) or absence (−) of 50 μM ivermectin in 22RV1 cells. J GSEA showed the inactivation of FOXA1 induced genes in 22RV1 cells after the ivermectin treatment. K RT-qPCR analysis of FL-AR and ARv7 in 22RV1 cells treated with ivermectin for 48 h. L ChIP–qPCR analysis for FOXA1 and FAIRE–qPCR analysis of chromatin accessibility at KLK3 and E2F1 in 22RV1 cells treated with ivermectin.

    Article Snippet: The samples were analyzed by immunoblotting with primary antibodies to: PARP (Cell Signaling Technology Cat# 9532, 1:1000), cleaved-caspase 3 (Cell Signaling Technology, Cat# 9664, 1:1000), γH2A.X (Cell Signaling Technology Cat# 2577, 1:1000), AR (Santa Cruz Biotechnology Cat# sc-7305, 1:1000), PSA (Cell Signaling Technology Cat# 5365, 1:1000), UBE2C (Cell Signaling Technology Cat# 14234, 1:200), E2F1 (Cell Signaling Technology Cat# 3742, 1:1000), FOXA1 (Cell Signaling Technology Cat# 53528, 1:1000), Ku70 (Cell Signaling Technology Cat# 4588, 1:1000), Ku80 (Cell Signaling Technology Cat# 2180, 1:1000), BRCA1 (Cell Signaling Technology Cat# 9009, 1:1000), Rad51 (Cell Signaling Technology Cat# 8875, 1:1000), Lamin B (Cell Signaling Technology Cat# 13435, 1:1000), and GAPDH (Santa Cruz Biotechnology Cat# sc-47724, 1:1000).

    Techniques: Quantitative RT-PCR, Western Blot, Expressing, Transfection

    In PCa, ivermectin could target FOXA1 and Ku70/Ku80. The interaction of ivermectin and FOXA1 reduced the chromatin accessibility of AR signaling and E2F1, leading to cell cycle arrest and inhibiting cell proliferation. The interaction of ivermectin and Ku70/Ku80 block the recruitment of Ku70/Ku80 to DSB sites. Cooperating with the downregulation of AR regulated homologous recombination repair genes, BRCA1 and Rad51, ivermectin increased intracellular DNA damage level and triggered synthetic lethality.

    Journal: Cell Death & Disease

    Article Title: Integrated analysis reveals FOXA1 and Ku70/Ku80 as targets of ivermectin in prostate cancer

    doi: 10.1038/s41419-022-05182-0

    Figure Lengend Snippet: In PCa, ivermectin could target FOXA1 and Ku70/Ku80. The interaction of ivermectin and FOXA1 reduced the chromatin accessibility of AR signaling and E2F1, leading to cell cycle arrest and inhibiting cell proliferation. The interaction of ivermectin and Ku70/Ku80 block the recruitment of Ku70/Ku80 to DSB sites. Cooperating with the downregulation of AR regulated homologous recombination repair genes, BRCA1 and Rad51, ivermectin increased intracellular DNA damage level and triggered synthetic lethality.

    Article Snippet: The samples were analyzed by immunoblotting with primary antibodies to: PARP (Cell Signaling Technology Cat# 9532, 1:1000), cleaved-caspase 3 (Cell Signaling Technology, Cat# 9664, 1:1000), γH2A.X (Cell Signaling Technology Cat# 2577, 1:1000), AR (Santa Cruz Biotechnology Cat# sc-7305, 1:1000), PSA (Cell Signaling Technology Cat# 5365, 1:1000), UBE2C (Cell Signaling Technology Cat# 14234, 1:200), E2F1 (Cell Signaling Technology Cat# 3742, 1:1000), FOXA1 (Cell Signaling Technology Cat# 53528, 1:1000), Ku70 (Cell Signaling Technology Cat# 4588, 1:1000), Ku80 (Cell Signaling Technology Cat# 2180, 1:1000), BRCA1 (Cell Signaling Technology Cat# 9009, 1:1000), Rad51 (Cell Signaling Technology Cat# 8875, 1:1000), Lamin B (Cell Signaling Technology Cat# 13435, 1:1000), and GAPDH (Santa Cruz Biotechnology Cat# sc-47724, 1:1000).

    Techniques: Blocking Assay, Homologous Recombination