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  • dmso  (ATCC)
    99
    ATCC dmso
    Dmso, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 619 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher pluronic f 127
    Pluronic F 127, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 5795 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dimethyl sulfoxide dmso
    Dimethyl Sulfoxide Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 20263 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dmso
    Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 55308 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher to pro 3 iodide 642 661 1 mm solution in dmso
    To Pro 3 Iodide 642 661 1 Mm Solution In Dmso, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2169 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher sytox green
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    Thermo Fisher sybr gold nucleic acid gel stain
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    92
    Fisher Scientific dmso
    Pilot screening using the BODIPY FL VH032 (5)-mediated VHL TR-FRET binding assay in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM <t>GST–VCB,</t> and 2 nM Tb-anti-GST at the 90-min incubation time point. (A) Plate-by-plate negative <t>(DMSO)</t> and positive (VH298, 2 , 30 μM) control performance. (B) Screening scatterplot of plate Z -prime. (C) Screening scatterplot of activity values, where the positive control is VH298 ( 2 , 30 μM, green dots, 100% inhibition), the negative control is DMSO (red dots, 0% inhibition), active compounds (blue dots) are chemicals with % inhibition ≥30% with the 30% activity cutoff defined by the black dotted line, and inactive compounds (black dots) are chemicals with % inhibition
    Dmso, supplied by Fisher Scientific, used in various techniques. Bioz Stars score: 92/100, based on 1380 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Merck KGaA dmso
    HSulf-1 in combination with <t>palbociclib</t> exerts synergistic antitumor effects by inducing G 1 /S stage cell cycle arrest and by inhibiting migration, invasion and epithelial-mesenchymal transition in vitro . (A-E) Hs578T and MDA-MB-231 cells following transient transfection with the control vector or HSulf-1 overexpression plasmids and incubation with 500 nM palbociclib or <t>DMSO</t> for 72 h. (A) Cell cycle analysis. G 1 and S phases of pcDNA3.0-Vector+Vehicle vs. pcDNA3.0-Vector+Palbociclib; G 1 and G 2 phases of pcDNA3.0-Vector+Vehicle vs. pcDNA3.0-HSulf-1+Vehicle; G 1 phase of pcDNA3.0-Vector+Palbociclib vs. pcDNA3.0-HSulf-1+Palbociclib; G 1 phase of pcDNA3.0-HSulf-1+Vehicle vs. pcDNA3.0-HSulf-1+Palbociclib. (B) Apoptosis analysis. (C) Wound healing assays. (D) Transwell assays. Magnification, x100. (E) Western blotting was performed to measure E-cadherin, vimentin and N-cadherin expression, with β-actin used as an internal control. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P
    Dmso, supplied by Merck KGaA, used in various techniques. Bioz Stars score: 92/100, based on 1546 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Merck KGaA dimethyl sulfoxide dmso
    In vivo activity of <t>EN4</t> in a mouse tissue cage model. The numbers of planktonic SA113 (A) and the viability of leukocytes (B) present in tissue cage fluid from cages treated with 100 or 250 μg of EN4 or 30 μg of daptomycin (DAP) and infected with 4 × 10 3 CFU of SA113 were investigated. Open circles indicate control mice injected with 1% <t>DMSO-PBS.</t> The values shown are means ± the SDs.
    Dimethyl Sulfoxide Dmso, supplied by Merck KGaA, used in various techniques. Bioz Stars score: 92/100, based on 813 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Fisher Scientific dimethyl sulfoxide dmso
    Analogs of <t>honokiol</t> do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle <t>(DMSO)</t> and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled
    Dimethyl Sulfoxide Dmso, supplied by Fisher Scientific, used in various techniques. Bioz Stars score: 92/100, based on 730 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Merck & Co dmso
    Dose-dependent antifungal effect of <t>L1</t> against Botrytis cinerea B05.10 at 4 °C. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in <t>DMSO</t> (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.
    Dmso, supplied by Merck & Co, used in various techniques. Bioz Stars score: 92/100, based on 970 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore hek293 cells
    DWORF activates SERCA2a in the absence of PLB. (A) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). (B) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF-P15/W22 (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). GFP-SERCA2a was stably expressed in <t>HEK293</t> cells, then 20 μg of DNA per dish (for unlabeled DWORF or unlabeled mutant DWORF) was transiently transfected into the cells. Ca-ATPase assays were performed using cell homogenates as described in Methods. Activity was normalized to the control (SERCA2a only), since DWORF had no significant effect on V max . (n=3)
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    Millipore dimethyl sulfoxide
    DWORF activates SERCA2a in the absence of PLB. (A) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). (B) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF-P15/W22 (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). GFP-SERCA2a was stably expressed in <t>HEK293</t> cells, then 20 μg of DNA per dish (for unlabeled DWORF or unlabeled mutant DWORF) was transiently transfected into the cells. Ca-ATPase assays were performed using cell homogenates as described in Methods. Activity was normalized to the control (SERCA2a only), since DWORF had no significant effect on V max . (n=3)
    Dimethyl Sulfoxide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 27840 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Carl Roth GmbH dmso
    Treatment with HIT and <t>SAHA</t> results in a reduced proliferation in osteosarcoma xenografts. Histological analysis of proliferation in tumors treated with <t>DMSO</t> (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The proliferation rate was significantly reduced in tumors after treatment with SAHA and HIT at all investigation time points. HIT only treatment led to a significantly lower proliferation rate 8 and 24 days after irradiation compared to the control group. SAHA only treatment had no significant effect on tumor proliferation
    Dmso, supplied by Carl Roth GmbH, used in various techniques. Bioz Stars score: 92/100, based on 622 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Amresco dmso
    <t>Gimatecan</t> Inhibited HCC Cell Growth in Vitro . Different HCC cell lines were cultured and incubated with various concentrations of gimatecan at 37 ºC for 72 hours. The percentages of viable cells were measured at the end the incubation period. 100% refers to the number of cells after 72 hours of incubation in the presence of the vehicle control (0.1% <t>DMSO).</t>
    Dmso, supplied by Amresco, used in various techniques. Bioz Stars score: 92/100, based on 499 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    FUJIFILM dmso
    Analysis of RelA-dependent signaling in RelA-sfGFP-S4D-KI cells. a Quantitative RT-PCR for the expression of TNF-α-induced genes. Parental or RelA-sfGFP-S4D-KI cells were pretreated with <t>DMSO</t> or pomalidomide (Po) for 24 h. Then, the cells were stimulated with 20 ng/ml TNF-α for 1 h, and the expression of IκBα or IL-6 was measured by quantitative RT-PCR. The mRNA expression in untreated parental HeLa cells was set to 1.0. b , c Pomalidomide causes TNF-α-induced cell death. Parental and RelA-sfGFP-S4D-KI cells pretreated with DMSO or Po for 12 h were stimulated with 20 ng/ml TNF-α for 12 h, and the viability was measured by MTS assay ( b ) or trypan blue staining ( c ). d Immunoblot analysis of pomalidomide-dependent TNF-α-induced cell death. Parental and RelA-sfGFP-S4D-KI cells pretreated with pomalidomide for 12 h were stimulated with 50 ng/ml TNF-α for the indicated times, and effectors of cell death were analyzed by immunoblot. e , f <t>zVAD-FMK</t> treatment rescued TNF-α-induced cell death in pomalidomide-treated KI cells. Parental and RelA-sfGFP-S4D-KI cells pretreated with 10 µM Po for 12 h were then treated with DMSO or 10 µM zVAD-FMK. After 2 h of zVAD-FMK treatment, the cells were stimulated with 50 ng/ml TNF-α for 12 h and the viability was measured by trypan blue staining ( e ), or effectors of apoptosis were analyzed by immunoblot ( f ). Error bars in a – c and e represent the mean ± SD ( n = 3), and P values were calculated by one-way ANOVA with Tukey’s post-hoc tests (NS not significant; * P
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    Merck & Co dimethyl sulfoxide dmso
    The combination of <t>LCRF‐0006</t> and bortezomib synergistically increases MM tumor response in vivo. A, Schematic representation of treatment regimen. C57BL/KaLwRij mice with established MM (day 14 post‐5TGM1 cell injection) were treated with LCRF‐0006 (100 mg/kg) and bortezomib (0.5 mg/kg) (LCRF‐0006 + bortezomib), LCRF‐0006 and <t>DMSO</t> vehicle (LCRF‐0006), 2‐HP‐b‐CD vehicle and bortezomib (bortezomib) or 2‐HP‐b‐CD and DMSO vehicles (vehicles) i.p., as per treatment regimen. B, Tumor burden was assessed at day 14, 21 and 28 by BLI. Graph depicts mean ± SEM. n = 5‐6 mice/treatment group. *** P
    Dimethyl Sulfoxide Dmso, supplied by Merck & Co, used in various techniques. Bioz Stars score: 92/100, based on 381 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    N/A
    Standard format Plasmid sent in bacteria as agar stab
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    Standard format Plasmid sent in bacteria as agar stab
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    Image Search Results


    Pilot screening using the BODIPY FL VH032 (5)-mediated VHL TR-FRET binding assay in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM GST–VCB, and 2 nM Tb-anti-GST at the 90-min incubation time point. (A) Plate-by-plate negative (DMSO) and positive (VH298, 2 , 30 μM) control performance. (B) Screening scatterplot of plate Z -prime. (C) Screening scatterplot of activity values, where the positive control is VH298 ( 2 , 30 μM, green dots, 100% inhibition), the negative control is DMSO (red dots, 0% inhibition), active compounds (blue dots) are chemicals with % inhibition ≥30% with the 30% activity cutoff defined by the black dotted line, and inactive compounds (black dots) are chemicals with % inhibition

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: Pilot screening using the BODIPY FL VH032 (5)-mediated VHL TR-FRET binding assay in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM GST–VCB, and 2 nM Tb-anti-GST at the 90-min incubation time point. (A) Plate-by-plate negative (DMSO) and positive (VH298, 2 , 30 μM) control performance. (B) Screening scatterplot of plate Z -prime. (C) Screening scatterplot of activity values, where the positive control is VH298 ( 2 , 30 μM, green dots, 100% inhibition), the negative control is DMSO (red dots, 0% inhibition), active compounds (blue dots) are chemicals with % inhibition ≥30% with the 30% activity cutoff defined by the black dotted line, and inactive compounds (black dots) are chemicals with % inhibition

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: Binding Assay, Incubation, Activity Assay, Positive Control, Inhibition, Negative Control

    TR-FRET binding affinity of BODIPY FL VH032 ( 5 , 1 to 2 dilutions, at an optimized concentration range of 0.06–500 nM) to GST–VCB. (A) Binding interaction of BODIPY FL VH032 ( 5 ) to 2 nM Tb-anti-GST in the presence of 2 nM GST–VCB or in the absence of GST–VCB at the designated incubation times. “+” and “–” (after each time point) represent “with GST–VCB” and “without GST–VCB”, respectively. The TR-FRET signals were expressed as relative TR-FRET units (RTU), which were calculated using 10,000 × 520 nm/490 nm. (B) Binding interaction of BODIPY FL VH032 ( 5 ) and 2 nM Tb-anti-GST, with 2 nM GST–VCB, 2 nM GST–VCB + VH298 ( 2 , 30 μM), or without GST–VCB + DMSO at the 90-min incubation time. (C) Fold changes in the TR-FRET signals of BODIPY FL VH032 ( 5 ) with (2 nM GST–VCB + DMSO) to (2 nM GST–VCB + VH298) ( 2 , 30 μM) (blue curve) or to (without GST–VCB + DMSO) (red curve).

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: TR-FRET binding affinity of BODIPY FL VH032 ( 5 , 1 to 2 dilutions, at an optimized concentration range of 0.06–500 nM) to GST–VCB. (A) Binding interaction of BODIPY FL VH032 ( 5 ) to 2 nM Tb-anti-GST in the presence of 2 nM GST–VCB or in the absence of GST–VCB at the designated incubation times. “+” and “–” (after each time point) represent “with GST–VCB” and “without GST–VCB”, respectively. The TR-FRET signals were expressed as relative TR-FRET units (RTU), which were calculated using 10,000 × 520 nm/490 nm. (B) Binding interaction of BODIPY FL VH032 ( 5 ) and 2 nM Tb-anti-GST, with 2 nM GST–VCB, 2 nM GST–VCB + VH298 ( 2 , 30 μM), or without GST–VCB + DMSO at the 90-min incubation time. (C) Fold changes in the TR-FRET signals of BODIPY FL VH032 ( 5 ) with (2 nM GST–VCB + DMSO) to (2 nM GST–VCB + VH298) ( 2 , 30 μM) (blue curve) or to (without GST–VCB + DMSO) (red curve).

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: Binding Assay, Concentration Assay, Incubation

    DMSO tolerance of the BODIPY FL VH032 ( 5 )-mediated VHL TR-FRET assay in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM GST–VCB, and 2 nM Tb-anti-GST at the 90-min incubation time point. (A) RTU of the negative control DMSO or the positive control VH298 ( 2 , 30 μM) in the presence of 0.2, 0.5, 1, 2, 5, or 10% DMSO. (B) % RTU (% RTU of 0.2% DMSO was set as 100%) of the negative control DMSO or the positive control VH298 ( 2 , 30 μM) in the presence of 0.2, 0.5, 1, 2, 5, or 10% DMSO. (C) Dose–response inhibition curves of VH298 ( 2 , 1–3 dilutions; in the concentration range of 2.1 pM to 30 μM) in the presence of indicated DMSO concentrations.

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: DMSO tolerance of the BODIPY FL VH032 ( 5 )-mediated VHL TR-FRET assay in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM GST–VCB, and 2 nM Tb-anti-GST at the 90-min incubation time point. (A) RTU of the negative control DMSO or the positive control VH298 ( 2 , 30 μM) in the presence of 0.2, 0.5, 1, 2, 5, or 10% DMSO. (B) % RTU (% RTU of 0.2% DMSO was set as 100%) of the negative control DMSO or the positive control VH298 ( 2 , 30 μM) in the presence of 0.2, 0.5, 1, 2, 5, or 10% DMSO. (C) Dose–response inhibition curves of VH298 ( 2 , 1–3 dilutions; in the concentration range of 2.1 pM to 30 μM) in the presence of indicated DMSO concentrations.

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: Incubation, Negative Control, Positive Control, Inhibition, Concentration Assay

    Activities of controls and selected VHL or non-VHL ligands in the BODIPY FL VH032 ( 5 )-mediated VHL FP assay with 10 nM BODIPY FL VH032 ( 5 ) and 100 nM GST–VCB at a 90-min incubation time. (A) FP assay signals of DMSO and VH298 ( 2 , 30 μM). (B) FP signal fold change from VH298 ( 2 , 30 μM) of DMSO and VH298 ( 2 , 30 μM). (C) FP dose–response curves of the VHL ligands VH032 ( 1 ), VH298 ( 2 ), VH032 amine ( 6 ), Me-VH032 amine ( 7 ), BOC-VH032 ( 8 ), and VH032 phenol ( 9 ). (D) FP dose–response curves of VH298 ( 2 ), MZ1 ( 3 ), and VH032-PEG4-amine ( 10 ) and of the non-VHL ligands (+)-JQ1 ( 4 ), thalidomide-4′-oxyacetamido-alkylC4-amine ( 11 ), and dBET1 ( 12 ).

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: Activities of controls and selected VHL or non-VHL ligands in the BODIPY FL VH032 ( 5 )-mediated VHL FP assay with 10 nM BODIPY FL VH032 ( 5 ) and 100 nM GST–VCB at a 90-min incubation time. (A) FP assay signals of DMSO and VH298 ( 2 , 30 μM). (B) FP signal fold change from VH298 ( 2 , 30 μM) of DMSO and VH298 ( 2 , 30 μM). (C) FP dose–response curves of the VHL ligands VH032 ( 1 ), VH298 ( 2 ), VH032 amine ( 6 ), Me-VH032 amine ( 7 ), BOC-VH032 ( 8 ), and VH032 phenol ( 9 ). (D) FP dose–response curves of VH298 ( 2 ), MZ1 ( 3 ), and VH032-PEG4-amine ( 10 ) and of the non-VHL ligands (+)-JQ1 ( 4 ), thalidomide-4′-oxyacetamido-alkylC4-amine ( 11 ), and dBET1 ( 12 ).

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: FP Assay, Incubation

    Determination of the binding affinity of BODIPY FL VH032 ( 5 , 10 nM) to GST–VCB in an FP assay with GST–VCB (1 to 2 dilutions; in the optimal concentration range of 0.03–1000 nM) + DMSO (total interactions), GST–VCB (1 to 2 dilutions; in the optimal concentration range of 0.03–1000 nM) + VH298 ( 2 , 30 μM) (i.e., GST–VCB-mediated nonspecific interactions), or DMSO without GST–VCB (i.e., background interactions) at a 90-min incubation time.

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: Determination of the binding affinity of BODIPY FL VH032 ( 5 , 10 nM) to GST–VCB in an FP assay with GST–VCB (1 to 2 dilutions; in the optimal concentration range of 0.03–1000 nM) + DMSO (total interactions), GST–VCB (1 to 2 dilutions; in the optimal concentration range of 0.03–1000 nM) + VH298 ( 2 , 30 μM) (i.e., GST–VCB-mediated nonspecific interactions), or DMSO without GST–VCB (i.e., background interactions) at a 90-min incubation time.

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: Binding Assay, FP Assay, Concentration Assay, Incubation

    Dose–response curves of a panel of VHL ligands and non-VHL ligands in the presence of BODIPY FL VH032 ( 5 , 4 nM), 2 nM GST–VCB, and 2 nM Tb-anti-GST at a 90-min incubation time. Ligand-relative TR-FRET units (RTUs) at their individual concentrations were normalized to that of VH298 ( 2 , 30 μM, positive control, 100% inhibition) and DMSO (negative control, 0% inhibition) and fitted to a sigmoidal equation with GraphPad PRISM to derive the IC 50 values, if applicable. The K i values were calculated with the Cheng–Prusoff equation. 35 (A) Dose–response curves of VHL ligands VH032 ( 1 ), VH298 ( 2 ), VH032 amine ( 6 ), Me-VH032 amine ( 7 ), BOC-VH032 ( 8 ), and VH032 phenol ( 9 ). (B) Dose–response curves of VHL ligands VH298 ( 2 ), MZ1 ( 3 ), and VH032-PEG4-amine ( 10 ) and of non-VHL ligands (+)-JQ1 ( 4 ), thalidomide-4′-oxyacetamido-alkylC4-amine ( 11 ), and dBET1 ( 12 ).

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: Dose–response curves of a panel of VHL ligands and non-VHL ligands in the presence of BODIPY FL VH032 ( 5 , 4 nM), 2 nM GST–VCB, and 2 nM Tb-anti-GST at a 90-min incubation time. Ligand-relative TR-FRET units (RTUs) at their individual concentrations were normalized to that of VH298 ( 2 , 30 μM, positive control, 100% inhibition) and DMSO (negative control, 0% inhibition) and fitted to a sigmoidal equation with GraphPad PRISM to derive the IC 50 values, if applicable. The K i values were calculated with the Cheng–Prusoff equation. 35 (A) Dose–response curves of VHL ligands VH032 ( 1 ), VH298 ( 2 ), VH032 amine ( 6 ), Me-VH032 amine ( 7 ), BOC-VH032 ( 8 ), and VH032 phenol ( 9 ). (B) Dose–response curves of VHL ligands VH298 ( 2 ), MZ1 ( 3 ), and VH032-PEG4-amine ( 10 ) and of non-VHL ligands (+)-JQ1 ( 4 ), thalidomide-4′-oxyacetamido-alkylC4-amine ( 11 ), and dBET1 ( 12 ).

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: Incubation, Positive Control, Inhibition, Negative Control

    Signal stability of the BODIPY FL VH032 ( 5 )-based VHL TR-FRET assay. (A) TR-FRET interaction of 4 nM BODIPY FL VH032 ( 5 ) and 2 nM Tb-anti-GST with 2 nM GST–VCB + DMSO (negative control), 2 nM GST–VCB + VH298 ( 2 , 30 μM) (positive control), or without GST–VCB + DMSO (background control) at specified incubation time points. (B) TR-FRET signal-fold change to 2 nM GST–VCB + VH298 ( 2 , 30 μM) of 2 nM GST–VCB + DMSO (negative control), 2 nM GST–VCB + VH298 ( 2 , 30 μM) (positive control), or without GST–VCB + DMSO (background control) in the presence of 4 nM BODIPY FL VH032 ( 5 ) and 2 nM Tb-anti-GST at specified incubation time points. (C) Dose–response inhibition curves of VH298 ( 2 , 1–3 dilutions, in the concentration range of 2.1 pM to 30 μM) at specified incubation time points in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM GST–VCB, and 2 nM Tb-anti-GST.

    Journal: ACS Omega

    Article Title: Development of BODIPY FL VH032 as a High-Affinity and Selective von Hippel–Lindau E3 Ligase Fluorescent Probe and Its Application in a Time-Resolved Fluorescence Resonance Energy-Transfer Assay

    doi: 10.1021/acsomega.0c05221

    Figure Lengend Snippet: Signal stability of the BODIPY FL VH032 ( 5 )-based VHL TR-FRET assay. (A) TR-FRET interaction of 4 nM BODIPY FL VH032 ( 5 ) and 2 nM Tb-anti-GST with 2 nM GST–VCB + DMSO (negative control), 2 nM GST–VCB + VH298 ( 2 , 30 μM) (positive control), or without GST–VCB + DMSO (background control) at specified incubation time points. (B) TR-FRET signal-fold change to 2 nM GST–VCB + VH298 ( 2 , 30 μM) of 2 nM GST–VCB + DMSO (negative control), 2 nM GST–VCB + VH298 ( 2 , 30 μM) (positive control), or without GST–VCB + DMSO (background control) in the presence of 4 nM BODIPY FL VH032 ( 5 ) and 2 nM Tb-anti-GST at specified incubation time points. (C) Dose–response inhibition curves of VH298 ( 2 , 1–3 dilutions, in the concentration range of 2.1 pM to 30 μM) at specified incubation time points in the presence of 4 nM BODIPY FL VH032 ( 5 ), 2 nM GST–VCB, and 2 nM Tb-anti-GST.

    Article Snippet: Biology The Tb-anti-GST antibody, 1,4-dithiothreitol (DTT, 1 M), Tris (1 M, pH 7.5), and DMSO were purchased from Fisher Scientific (Pittsburgh, PA).

    Techniques: Negative Control, Positive Control, Incubation, Inhibition, Concentration Assay

    HSulf-1 in combination with palbociclib exerts synergistic antitumor effects by inducing G 1 /S stage cell cycle arrest and by inhibiting migration, invasion and epithelial-mesenchymal transition in vitro . (A-E) Hs578T and MDA-MB-231 cells following transient transfection with the control vector or HSulf-1 overexpression plasmids and incubation with 500 nM palbociclib or DMSO for 72 h. (A) Cell cycle analysis. G 1 and S phases of pcDNA3.0-Vector+Vehicle vs. pcDNA3.0-Vector+Palbociclib; G 1 and G 2 phases of pcDNA3.0-Vector+Vehicle vs. pcDNA3.0-HSulf-1+Vehicle; G 1 phase of pcDNA3.0-Vector+Palbociclib vs. pcDNA3.0-HSulf-1+Palbociclib; G 1 phase of pcDNA3.0-HSulf-1+Vehicle vs. pcDNA3.0-HSulf-1+Palbociclib. (B) Apoptosis analysis. (C) Wound healing assays. (D) Transwell assays. Magnification, x100. (E) Western blotting was performed to measure E-cadherin, vimentin and N-cadherin expression, with β-actin used as an internal control. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P

    Journal: International Journal of Oncology

    Article Title: HSulf-1 and palbociclib exert synergistic antitumor effects on RB-positive triple-negative breast cancer

    doi: 10.3892/ijo.2020.5057

    Figure Lengend Snippet: HSulf-1 in combination with palbociclib exerts synergistic antitumor effects by inducing G 1 /S stage cell cycle arrest and by inhibiting migration, invasion and epithelial-mesenchymal transition in vitro . (A-E) Hs578T and MDA-MB-231 cells following transient transfection with the control vector or HSulf-1 overexpression plasmids and incubation with 500 nM palbociclib or DMSO for 72 h. (A) Cell cycle analysis. G 1 and S phases of pcDNA3.0-Vector+Vehicle vs. pcDNA3.0-Vector+Palbociclib; G 1 and G 2 phases of pcDNA3.0-Vector+Vehicle vs. pcDNA3.0-HSulf-1+Vehicle; G 1 phase of pcDNA3.0-Vector+Palbociclib vs. pcDNA3.0-HSulf-1+Palbociclib; G 1 phase of pcDNA3.0-HSulf-1+Vehicle vs. pcDNA3.0-HSulf-1+Palbociclib. (B) Apoptosis analysis. (C) Wound healing assays. (D) Transwell assays. Magnification, x100. (E) Western blotting was performed to measure E-cadherin, vimentin and N-cadherin expression, with β-actin used as an internal control. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P

    Article Snippet: Palbociclib (Sigma-Aldrich; Merck KGaA) was dissolved in DMSO, following which the cells were treated with 500 nM palboci-clib for 72 h at 37°C.

    Techniques: Migration, In Vitro, Multiple Displacement Amplification, Transfection, Plasmid Preparation, Over Expression, Incubation, Cell Cycle Assay, Western Blot, Expressing, Derivative Assay

    HSulf-1 exhibit synergy with palbociclib to exert antiproliferative effects on breast cancer cells by reducing palbociclib-induced cyclin D1 accumulation. (A) Western blotting was performed to measure the protein levels of cyclin D1, p-RB and total RB in Hs578T and MDA-MB-231 cells following transient transfection with the control vector or HSulf-1 overexpression plasmids and incubation with 500 nM palbociclib or DMSO. β-Actin was used as an internal control. (B) Reverse transcription-quantitative PCR was performed to measure mRNA level of cyclin D1 in MDA-MB-231 cells following transient transfection with the control vector or cyclin D1 overexpression plasmids. GAPDH was used as an internal control. (C) Cell Counting Kit-8 and (D) colony formation assays were performed to evaluate cell viability in MDA-MB-231 cells following incubation with 500 nM palbociclib for 14 days, overexpression of HSulf-1, overexpression of HSulf-1 followed by incubation with 500 nM palbociclib for 14 days or co-transfection of plasmids encoding HSulf-1 and cyclin D1 followed by exposure to palbociclib for 14 days. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P

    Journal: International Journal of Oncology

    Article Title: HSulf-1 and palbociclib exert synergistic antitumor effects on RB-positive triple-negative breast cancer

    doi: 10.3892/ijo.2020.5057

    Figure Lengend Snippet: HSulf-1 exhibit synergy with palbociclib to exert antiproliferative effects on breast cancer cells by reducing palbociclib-induced cyclin D1 accumulation. (A) Western blotting was performed to measure the protein levels of cyclin D1, p-RB and total RB in Hs578T and MDA-MB-231 cells following transient transfection with the control vector or HSulf-1 overexpression plasmids and incubation with 500 nM palbociclib or DMSO. β-Actin was used as an internal control. (B) Reverse transcription-quantitative PCR was performed to measure mRNA level of cyclin D1 in MDA-MB-231 cells following transient transfection with the control vector or cyclin D1 overexpression plasmids. GAPDH was used as an internal control. (C) Cell Counting Kit-8 and (D) colony formation assays were performed to evaluate cell viability in MDA-MB-231 cells following incubation with 500 nM palbociclib for 14 days, overexpression of HSulf-1, overexpression of HSulf-1 followed by incubation with 500 nM palbociclib for 14 days or co-transfection of plasmids encoding HSulf-1 and cyclin D1 followed by exposure to palbociclib for 14 days. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P

    Article Snippet: Palbociclib (Sigma-Aldrich; Merck KGaA) was dissolved in DMSO, following which the cells were treated with 500 nM palboci-clib for 72 h at 37°C.

    Techniques: Western Blot, Multiple Displacement Amplification, Transfection, Plasmid Preparation, Over Expression, Incubation, Real-time Polymerase Chain Reaction, Cell Counting, Cotransfection, Derivative Assay

    HSulf-1 inhibits cell proliferation and when combined with palbociclib, exerts antiproliferative effects on RB-positive TNBC cells in a synergistic manner both in vitro and in vivo . (A) Hs578T and MDA-MB-231 cells were transiently transfected with the control vector or HSulf-1 overexpression plasmids and incubated for 6 h at 37°C then incubated with fresh complete medium cantaining 500 nM palbociclib or DMSO for 0, 24, 48 and 72 h. Subsequently, Cell Counting Kit-8 assays were performed to assess cell viability. (B) Hs578T and MDA-MB-231 cells were transiently transfected with the control vector or HSulf-1 overexpression plasmids and incubated for 6 h at 37°C then incubated with fresh complete medium containing 500 nM palbociclib or DMSO for up to 14 days for colony formation assay. (C) Representative images of the mouse xenograft models and tumors extracted from the four treatment groups after euthanasia. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P

    Journal: International Journal of Oncology

    Article Title: HSulf-1 and palbociclib exert synergistic antitumor effects on RB-positive triple-negative breast cancer

    doi: 10.3892/ijo.2020.5057

    Figure Lengend Snippet: HSulf-1 inhibits cell proliferation and when combined with palbociclib, exerts antiproliferative effects on RB-positive TNBC cells in a synergistic manner both in vitro and in vivo . (A) Hs578T and MDA-MB-231 cells were transiently transfected with the control vector or HSulf-1 overexpression plasmids and incubated for 6 h at 37°C then incubated with fresh complete medium cantaining 500 nM palbociclib or DMSO for 0, 24, 48 and 72 h. Subsequently, Cell Counting Kit-8 assays were performed to assess cell viability. (B) Hs578T and MDA-MB-231 cells were transiently transfected with the control vector or HSulf-1 overexpression plasmids and incubated for 6 h at 37°C then incubated with fresh complete medium containing 500 nM palbociclib or DMSO for up to 14 days for colony formation assay. (C) Representative images of the mouse xenograft models and tumors extracted from the four treatment groups after euthanasia. The results were derived from three independent experiments. Data are presented as the mean ± SD. * P

    Article Snippet: Palbociclib (Sigma-Aldrich; Merck KGaA) was dissolved in DMSO, following which the cells were treated with 500 nM palboci-clib for 72 h at 37°C.

    Techniques: In Vitro, In Vivo, Multiple Displacement Amplification, Transfection, Plasmid Preparation, Over Expression, Incubation, Cell Counting, Colony Assay, Derivative Assay

    In vivo activity of EN4 in a mouse tissue cage model. The numbers of planktonic SA113 (A) and the viability of leukocytes (B) present in tissue cage fluid from cages treated with 100 or 250 μg of EN4 or 30 μg of daptomycin (DAP) and infected with 4 × 10 3 CFU of SA113 were investigated. Open circles indicate control mice injected with 1% DMSO-PBS. The values shown are means ± the SDs.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Antimicrobial Properties of 8-Hydroxyserrulat-14-en-19-oic Acid for Treatment of Implant-Associated Infections

    doi: 10.1128/AAC.01735-12

    Figure Lengend Snippet: In vivo activity of EN4 in a mouse tissue cage model. The numbers of planktonic SA113 (A) and the viability of leukocytes (B) present in tissue cage fluid from cages treated with 100 or 250 μg of EN4 or 30 μg of daptomycin (DAP) and infected with 4 × 10 3 CFU of SA113 were investigated. Open circles indicate control mice injected with 1% DMSO-PBS. The values shown are means ± the SDs.

    Article Snippet: EN4 was dissolved in 1% dimethyl sulfoxide (DMSO) (Merck, Darmstadt, Germany) in phosphate-buffered saline (PBS) (Reagens, Basel, Switzerland) (DMSO-PBS) up to a concentration of 400 μg/ml.

    Techniques: In Vivo, Activity Assay, Infection, Mouse Assay, Injection

    Effect of EN4 on bacterial membrane integrity. (A) S. epidermidis 1457 was incubated for 10 min with EN4 or CHX at 100 μg/ml or 1% DMSO-PBS and subsequently double stained with propidium iodide (PI) and SYTO9 and analyzed by flow cytometry. The results of one representative experiment of three performed are shown. (B) Transmission electron microscopy images of ultrathin sections of WSPPA treated for 1 h with 1% DMSO-PBS as a control, EN4 at 100 or 200 μg/ml, or 250 μg of CHX/ml. Arrowheads indicate ultrastructural changes; the bars represent 200 nm. (C) WSPPA was treated for 10 min with EN4 or CHX at 100 μg/ml, followed by centrifugation and investigation of supernatants for the presence of ATP using luciferase reaction. Nisin (NIS) and ciprofloxacin (CIP) were used as a positive and a negative control, respectively, and 1% DMSO-PBS was used as an untreated control (Ctrl). The values shown are means ± the SDs of areas under the concentration-time curve calculated for the first 30 min of luciferase reaction from at least three independent experiments prepared in duplicates. Significant ATP leakage compared to results for the untreated control is indicated (*, P

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Antimicrobial Properties of 8-Hydroxyserrulat-14-en-19-oic Acid for Treatment of Implant-Associated Infections

    doi: 10.1128/AAC.01735-12

    Figure Lengend Snippet: Effect of EN4 on bacterial membrane integrity. (A) S. epidermidis 1457 was incubated for 10 min with EN4 or CHX at 100 μg/ml or 1% DMSO-PBS and subsequently double stained with propidium iodide (PI) and SYTO9 and analyzed by flow cytometry. The results of one representative experiment of three performed are shown. (B) Transmission electron microscopy images of ultrathin sections of WSPPA treated for 1 h with 1% DMSO-PBS as a control, EN4 at 100 or 200 μg/ml, or 250 μg of CHX/ml. Arrowheads indicate ultrastructural changes; the bars represent 200 nm. (C) WSPPA was treated for 10 min with EN4 or CHX at 100 μg/ml, followed by centrifugation and investigation of supernatants for the presence of ATP using luciferase reaction. Nisin (NIS) and ciprofloxacin (CIP) were used as a positive and a negative control, respectively, and 1% DMSO-PBS was used as an untreated control (Ctrl). The values shown are means ± the SDs of areas under the concentration-time curve calculated for the first 30 min of luciferase reaction from at least three independent experiments prepared in duplicates. Significant ATP leakage compared to results for the untreated control is indicated (*, P

    Article Snippet: EN4 was dissolved in 1% dimethyl sulfoxide (DMSO) (Merck, Darmstadt, Germany) in phosphate-buffered saline (PBS) (Reagens, Basel, Switzerland) (DMSO-PBS) up to a concentration of 400 μg/ml.

    Techniques: Incubation, Staining, Flow Cytometry, Cytometry, Transmission Assay, Electron Microscopy, Centrifugation, Luciferase, Negative Control, Concentration Assay

    Analogs of honokiol do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle (DMSO) and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled

    Journal: Cell Death & Disease

    Article Title: Honokiol is a FOXM1 antagonist

    doi: 10.1038/s41419-017-0156-7

    Figure Lengend Snippet: Analogs of honokiol do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle (DMSO) and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled

    Article Snippet: MG132 (EMD Millipore), thiostrepton (Sigma), honokiol, 2,2 dihydroxybiphenyl, 2 allylphenol, eugenol, and O -eugenol were dissolved in dimethyl sulfoxide (DMSO) (Fisher Scientific), N-acetyl-l-cysteine (NAC) (Sigma) in deionized water, and doxycycline (LKT Laboratories) in phosphate buffered saline (PBS).

    Techniques: Concentration Assay, Luciferase, Activity Assay, Nuclear Magnetic Resonance, Labeling, Recombinant

    Honokiol inhibits FOXM1 transactivation via binding a C3-luc cells were induced with doxycycline and treated with honokiol for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b The C3 cell line was treated with doxycycline and honokiol in the indicated concentrations for 24 h. Cells were collected and immunoblotting was performed with a FOXM1 specific antibody. β-actin was used as the loading control. c Representative EMSA image shows the inhibitory effect of honokiol on the formation of the FOXM1 DBD protein–DNA complex. d The C3 cell line was treated with doxycycline and honokiol as indicated for 24 h. Then, cells were processed for the ChIP experiments, as described in “Materials and methods”. Graph shows mean ± SEM of two independent ChIP experiments. e Saturation transfer difference (STD) NMR spectra to assess the binding of honokiol to FOXM1: (I) 2 mM of honokiol alone, (II) 150 ng of recombinant FOXM1 alone, (III) 2 mM honokiol with 150 ng of recombinant FOXM1. The chemical structure of honokiol is illustrated. STD signals arising from the aryl groups in honokiol are annotated, and signals from vehicle (DMSO) and water are labeled

    Journal: Cell Death & Disease

    Article Title: Honokiol is a FOXM1 antagonist

    doi: 10.1038/s41419-017-0156-7

    Figure Lengend Snippet: Honokiol inhibits FOXM1 transactivation via binding a C3-luc cells were induced with doxycycline and treated with honokiol for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b The C3 cell line was treated with doxycycline and honokiol in the indicated concentrations for 24 h. Cells were collected and immunoblotting was performed with a FOXM1 specific antibody. β-actin was used as the loading control. c Representative EMSA image shows the inhibitory effect of honokiol on the formation of the FOXM1 DBD protein–DNA complex. d The C3 cell line was treated with doxycycline and honokiol as indicated for 24 h. Then, cells were processed for the ChIP experiments, as described in “Materials and methods”. Graph shows mean ± SEM of two independent ChIP experiments. e Saturation transfer difference (STD) NMR spectra to assess the binding of honokiol to FOXM1: (I) 2 mM of honokiol alone, (II) 150 ng of recombinant FOXM1 alone, (III) 2 mM honokiol with 150 ng of recombinant FOXM1. The chemical structure of honokiol is illustrated. STD signals arising from the aryl groups in honokiol are annotated, and signals from vehicle (DMSO) and water are labeled

    Article Snippet: MG132 (EMD Millipore), thiostrepton (Sigma), honokiol, 2,2 dihydroxybiphenyl, 2 allylphenol, eugenol, and O -eugenol were dissolved in dimethyl sulfoxide (DMSO) (Fisher Scientific), N-acetyl-l-cysteine (NAC) (Sigma) in deionized water, and doxycycline (LKT Laboratories) in phosphate buffered saline (PBS).

    Techniques: Binding Assay, Luciferase, Activity Assay, Chromatin Immunoprecipitation, Nuclear Magnetic Resonance, Recombinant, Labeling

    Dose-dependent antifungal effect of L1 against Botrytis cinerea B05.10 at 4 °C. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Journal: Molecules

    Article Title: Structural Characterization, DFT Calculation, NCI, Scan-Rate Analysis and Antifungal Activity against Botrytis cinerea of (E)-2-{[(2-Aminopyridin-2-yl)imino]-methyl}-4,6-di-tert-butylphenol (Pyridine Schiff Base)

    doi: 10.3390/molecules25122741

    Figure Lengend Snippet: Dose-dependent antifungal effect of L1 against Botrytis cinerea B05.10 at 4 °C. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Article Snippet: To this end, Petri dishes (90 mm) with a final volume of 15 mL of PDA (Difco) containing 0 (negative control), 2, 4, 6, 8 or 10 ppm of L1 dissolved in DMSO (Merck) were used.

    Techniques: Inhibition, Concentration Assay

    Antifungal effect against Botrytis cinerea A1 (at 26 °C) exerted by L1 in a dose-dependent manner (12 days of incubation). Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (6 ppm) and compared with the commercial fungicide fenhexamid (6 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (0.045% v/v ). The figure shows a representative experiment, showing the fungal colony growth.

    Journal: Molecules

    Article Title: Structural Characterization, DFT Calculation, NCI, Scan-Rate Analysis and Antifungal Activity against Botrytis cinerea of (E)-2-{[(2-Aminopyridin-2-yl)imino]-methyl}-4,6-di-tert-butylphenol (Pyridine Schiff Base)

    doi: 10.3390/molecules25122741

    Figure Lengend Snippet: Antifungal effect against Botrytis cinerea A1 (at 26 °C) exerted by L1 in a dose-dependent manner (12 days of incubation). Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (6 ppm) and compared with the commercial fungicide fenhexamid (6 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (0.045% v/v ). The figure shows a representative experiment, showing the fungal colony growth.

    Article Snippet: To this end, Petri dishes (90 mm) with a final volume of 15 mL of PDA (Difco) containing 0 (negative control), 2, 4, 6, 8 or 10 ppm of L1 dissolved in DMSO (Merck) were used.

    Techniques: Incubation, Inhibition, Concentration Assay

    Dose-dependent antifungal effect of L1 against Botrytis cinerea A1 at 4 °C. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Journal: Molecules

    Article Title: Structural Characterization, DFT Calculation, NCI, Scan-Rate Analysis and Antifungal Activity against Botrytis cinerea of (E)-2-{[(2-Aminopyridin-2-yl)imino]-methyl}-4,6-di-tert-butylphenol (Pyridine Schiff Base)

    doi: 10.3390/molecules25122741

    Figure Lengend Snippet: Dose-dependent antifungal effect of L1 against Botrytis cinerea A1 at 4 °C. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Article Snippet: To this end, Petri dishes (90 mm) with a final volume of 15 mL of PDA (Difco) containing 0 (negative control), 2, 4, 6, 8 or 10 ppm of L1 dissolved in DMSO (Merck) were used.

    Techniques: Inhibition, Concentration Assay

    Antifungal effect against Botrytis cinerea A1 (at 26 °C) exerted by L1 in a dose-dependent manner. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Journal: Molecules

    Article Title: Structural Characterization, DFT Calculation, NCI, Scan-Rate Analysis and Antifungal Activity against Botrytis cinerea of (E)-2-{[(2-Aminopyridin-2-yl)imino]-methyl}-4,6-di-tert-butylphenol (Pyridine Schiff Base)

    doi: 10.3390/molecules25122741

    Figure Lengend Snippet: Antifungal effect against Botrytis cinerea A1 (at 26 °C) exerted by L1 in a dose-dependent manner. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Article Snippet: To this end, Petri dishes (90 mm) with a final volume of 15 mL of PDA (Difco) containing 0 (negative control), 2, 4, 6, 8 or 10 ppm of L1 dissolved in DMSO (Merck) were used.

    Techniques: Inhibition, Concentration Assay

    Antifungal effect against Botrytis cinerea B05.10 (at 26 °C) exerted by L1 in a dose-dependent manner. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Journal: Molecules

    Article Title: Structural Characterization, DFT Calculation, NCI, Scan-Rate Analysis and Antifungal Activity against Botrytis cinerea of (E)-2-{[(2-Aminopyridin-2-yl)imino]-methyl}-4,6-di-tert-butylphenol (Pyridine Schiff Base)

    doi: 10.3390/molecules25122741

    Figure Lengend Snippet: Antifungal effect against Botrytis cinerea B05.10 (at 26 °C) exerted by L1 in a dose-dependent manner. Inhibition of mycelial growth (by measuring the colony diameter) was observed in the presence of L1 (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm) and compared with the commercial fungicide fenhexamid (( A ): 0 ppm, ( B ): 2 ppm, ( C ): 4 ppm, ( D ): 6 ppm, ( E ): 8 ppm, ( F ): 10 ppm). Since both L1 and fenhexamid stock were dissolved in DMSO (vehicle), DMSO alone was also tested, adding the same concentration used with either L1 or fenhexamid in each case (( A ): 0% v/v , ( B ): 0.015% v/v , ( C ): 0.030% v/v , ( D ): 0.045% v/v , ( E ): 0.060% v/v , ( F ): 0.075% v/v ) (DMSO [eq]). In all cases, the experiments were performed in biological triplicate.

    Article Snippet: To this end, Petri dishes (90 mm) with a final volume of 15 mL of PDA (Difco) containing 0 (negative control), 2, 4, 6, 8 or 10 ppm of L1 dissolved in DMSO (Merck) were used.

    Techniques: Inhibition, Concentration Assay

    DWORF activates SERCA2a in the absence of PLB. (A) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). (B) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF-P15/W22 (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). GFP-SERCA2a was stably expressed in HEK293 cells, then 20 μg of DNA per dish (for unlabeled DWORF or unlabeled mutant DWORF) was transiently transfected into the cells. Ca-ATPase assays were performed using cell homogenates as described in Methods. Activity was normalized to the control (SERCA2a only), since DWORF had no significant effect on V max . (n=3)

    Journal: bioRxiv

    Article Title: The transmembrane domain of DWORF activates SERCA directly; P15 and W22 residues are essential

    doi: 10.1101/2020.09.18.303644

    Figure Lengend Snippet: DWORF activates SERCA2a in the absence of PLB. (A) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). (B) Ca-ATPase activity measured in homogenates of cells expressing GFP-SERCA2a and unlabeled DWORF-P15/W22 (black, pK Ca = 6.36 ± 0.04) or GFP-SERCA2a only (red, pK Ca = 6.08 ± 0.03). GFP-SERCA2a was stably expressed in HEK293 cells, then 20 μg of DNA per dish (for unlabeled DWORF or unlabeled mutant DWORF) was transiently transfected into the cells. Ca-ATPase assays were performed using cell homogenates as described in Methods. Activity was normalized to the control (SERCA2a only), since DWORF had no significant effect on V max . (n=3)

    Article Snippet: 24 hours before transfection, HEK293 cells were plated at 0.4 million cells per well in 6-well plates, or 2 million cells per dish in 10-centimeter dishes.

    Techniques: Activity Assay, Expressing, Stable Transfection, Mutagenesis, Transfection

    FRET-detected interaction of SERCA with the isolated TM domain of DWORF is similar to that with full-length DWORF. (A) FRET efficiency (E) in HEK293 cells stably expressing GFP-SERCA2a, vs. μg/dish of DNA transiently transfected into HEK293 cells expressing RFP-TM-DWORF (black) or full-length RFP-DWORF (red). (B) Fraction X DA of donors (GFP) transferring energy (bound) to acceptor (RFP) determined for samples in panel A. (C) FRET efficiency E for increasing amounts of TM-DWORF DNA transfected into a stable cell line expressing GFP-SERCA2a and RFP-PLB. (D) FRET efficiency plotted vs. DNA transfected per well. D. Fraction X DA of donors transferring energy (bound) to acceptors determined for samples in C.

    Journal: bioRxiv

    Article Title: The transmembrane domain of DWORF activates SERCA directly; P15 and W22 residues are essential

    doi: 10.1101/2020.09.18.303644

    Figure Lengend Snippet: FRET-detected interaction of SERCA with the isolated TM domain of DWORF is similar to that with full-length DWORF. (A) FRET efficiency (E) in HEK293 cells stably expressing GFP-SERCA2a, vs. μg/dish of DNA transiently transfected into HEK293 cells expressing RFP-TM-DWORF (black) or full-length RFP-DWORF (red). (B) Fraction X DA of donors (GFP) transferring energy (bound) to acceptor (RFP) determined for samples in panel A. (C) FRET efficiency E for increasing amounts of TM-DWORF DNA transfected into a stable cell line expressing GFP-SERCA2a and RFP-PLB. (D) FRET efficiency plotted vs. DNA transfected per well. D. Fraction X DA of donors transferring energy (bound) to acceptors determined for samples in C.

    Article Snippet: 24 hours before transfection, HEK293 cells were plated at 0.4 million cells per well in 6-well plates, or 2 million cells per dish in 10-centimeter dishes.

    Techniques: Isolation, Stable Transfection, Expressing, Transfection, Transferring

    RFP-DWORF binding to GFP-SERCA2a in live cells. (A) Confocal microscopy images of GFP-SERCA (donor) and RFP-DWORF (acceptor) fluorescence signals. (B) Fluorescence lifetime (FLT) traces measured from HEK293 cells stably expressing the donor without (green, F D (t) or with (red, F D+A (t)) transient expression of the acceptor. FLT data was analyzed as described in Methods to determine lifetimes (τ D and τ D+A ) and mole fractions X D and X DA . (C) FRET efficiency E (1-τ D+A /τ D ) vs. μg of acceptor DNA seeded per dish. (D) The fraction of SERCA2a containing bound DWORF (X DA ).

    Journal: bioRxiv

    Article Title: The transmembrane domain of DWORF activates SERCA directly; P15 and W22 residues are essential

    doi: 10.1101/2020.09.18.303644

    Figure Lengend Snippet: RFP-DWORF binding to GFP-SERCA2a in live cells. (A) Confocal microscopy images of GFP-SERCA (donor) and RFP-DWORF (acceptor) fluorescence signals. (B) Fluorescence lifetime (FLT) traces measured from HEK293 cells stably expressing the donor without (green, F D (t) or with (red, F D+A (t)) transient expression of the acceptor. FLT data was analyzed as described in Methods to determine lifetimes (τ D and τ D+A ) and mole fractions X D and X DA . (C) FRET efficiency E (1-τ D+A /τ D ) vs. μg of acceptor DNA seeded per dish. (D) The fraction of SERCA2a containing bound DWORF (X DA ).

    Article Snippet: 24 hours before transfection, HEK293 cells were plated at 0.4 million cells per well in 6-well plates, or 2 million cells per dish in 10-centimeter dishes.

    Techniques: Binding Assay, Confocal Microscopy, Fluorescence, Stable Transfection, Expressing

    RFP-DWORF activates SERCA2a directly in the absence of PLB. Transient transfections of 5μg of RFP-DWORF into HEK293 cells stably expressing GFP-SERCA2a. 48 hours after transfection, cells was harvested at 10 million per mL in homogenization buffer (0.5mM MgCl2, 10mM Tris-HCL ph 7.5, DNase I and protease inhibitor). Cells was then lysed by Tissumizer (Tekmar SDT-1810) and prepared in 96well plate for ATPase assay. n=3

    Journal: bioRxiv

    Article Title: The transmembrane domain of DWORF activates SERCA directly; P15 and W22 residues are essential

    doi: 10.1101/2020.09.18.303644

    Figure Lengend Snippet: RFP-DWORF activates SERCA2a directly in the absence of PLB. Transient transfections of 5μg of RFP-DWORF into HEK293 cells stably expressing GFP-SERCA2a. 48 hours after transfection, cells was harvested at 10 million per mL in homogenization buffer (0.5mM MgCl2, 10mM Tris-HCL ph 7.5, DNase I and protease inhibitor). Cells was then lysed by Tissumizer (Tekmar SDT-1810) and prepared in 96well plate for ATPase assay. n=3

    Article Snippet: 24 hours before transfection, HEK293 cells were plated at 0.4 million cells per well in 6-well plates, or 2 million cells per dish in 10-centimeter dishes.

    Techniques: Transfection, Stable Transfection, Expressing, Homogenization, Protease Inhibitor, ATPase Assay

    TM-DWORF can activate SERCA2a directly in the absence of PLB. Transient transfections of 5μg of TM-DWORF into HEK293 cells stably expressing GFP-SERCA2a. 48 hours after transfection, cells was harvested at 10 million per mL in homogenization buffer (0.5mM MgCl2, 10mM Tris-HCL ph 7.5, DNase I and protease inhibitor). Cells was then lysed by Tissumizer (Tekmar SDT-1810) and prepared in 96well plate for ATPase assay. n=3

    Journal: bioRxiv

    Article Title: The transmembrane domain of DWORF activates SERCA directly; P15 and W22 residues are essential

    doi: 10.1101/2020.09.18.303644

    Figure Lengend Snippet: TM-DWORF can activate SERCA2a directly in the absence of PLB. Transient transfections of 5μg of TM-DWORF into HEK293 cells stably expressing GFP-SERCA2a. 48 hours after transfection, cells was harvested at 10 million per mL in homogenization buffer (0.5mM MgCl2, 10mM Tris-HCL ph 7.5, DNase I and protease inhibitor). Cells was then lysed by Tissumizer (Tekmar SDT-1810) and prepared in 96well plate for ATPase assay. n=3

    Article Snippet: 24 hours before transfection, HEK293 cells were plated at 0.4 million cells per well in 6-well plates, or 2 million cells per dish in 10-centimeter dishes.

    Techniques: Transfection, Stable Transfection, Expressing, Homogenization, Protease Inhibitor, ATPase Assay

    Treatment with HIT and SAHA results in a reduced proliferation in osteosarcoma xenografts. Histological analysis of proliferation in tumors treated with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The proliferation rate was significantly reduced in tumors after treatment with SAHA and HIT at all investigation time points. HIT only treatment led to a significantly lower proliferation rate 8 and 24 days after irradiation compared to the control group. SAHA only treatment had no significant effect on tumor proliferation

    Journal: Radiation Oncology (London, England)

    Article Title: Histone deacetylase inhibition sensitizes osteosarcoma to heavy ion radiotherapy

    doi: 10.1186/s13014-015-0455-z

    Figure Lengend Snippet: Treatment with HIT and SAHA results in a reduced proliferation in osteosarcoma xenografts. Histological analysis of proliferation in tumors treated with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The proliferation rate was significantly reduced in tumors after treatment with SAHA and HIT at all investigation time points. HIT only treatment led to a significantly lower proliferation rate 8 and 24 days after irradiation compared to the control group. SAHA only treatment had no significant effect on tumor proliferation

    Article Snippet: SAHA was obtained from Alexis Biochemicals (Lörrach, Germany), DMSO from Carl Roth Biochemicals (Karlsruhe, Germany).

    Techniques: Irradiation

    Density of microvessels is significantly reduced in osteosarcoma after combination treatment. Xenografts were treated with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The density of microvessels was reduced in tumors of all treatment groups at all investigation time points. Tumors treated with HIT and SAHA showed a significant (*) lower vascularization compared to tumors treated with HIT from day 24 on and the lowest vascularization at all

    Journal: Radiation Oncology (London, England)

    Article Title: Histone deacetylase inhibition sensitizes osteosarcoma to heavy ion radiotherapy

    doi: 10.1186/s13014-015-0455-z

    Figure Lengend Snippet: Density of microvessels is significantly reduced in osteosarcoma after combination treatment. Xenografts were treated with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The density of microvessels was reduced in tumors of all treatment groups at all investigation time points. Tumors treated with HIT and SAHA showed a significant (*) lower vascularization compared to tumors treated with HIT from day 24 on and the lowest vascularization at all

    Article Snippet: SAHA was obtained from Alexis Biochemicals (Lörrach, Germany), DMSO from Carl Roth Biochemicals (Karlsruhe, Germany).

    Techniques: Irradiation

    Impairment of apoptosis and necrosis after HIT and SAHA treatment. Apoptosis and necrosis in osteosarcoma xenografts were analyzed after treatment with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The combination treatment lead to a significant (*) induction of apoptosis one day, 8 and 45 days after HIT compared to HIT only treatment. Apoptosis was also increased after SAHA only and HIT only 24 h, 8 and 45 days after irradiation but not as much as treatment with HIT and SAHA. Combination of HIT and SAHA lead to a significant (*) induction of necrosis on day 45. SAHA only and HIT only treatment resulted in a significantly higher rate of necrosis from day 24 on but not at earlier time points compared to the control groups

    Journal: Radiation Oncology (London, England)

    Article Title: Histone deacetylase inhibition sensitizes osteosarcoma to heavy ion radiotherapy

    doi: 10.1186/s13014-015-0455-z

    Figure Lengend Snippet: Impairment of apoptosis and necrosis after HIT and SAHA treatment. Apoptosis and necrosis in osteosarcoma xenografts were analyzed after treatment with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. The combination treatment lead to a significant (*) induction of apoptosis one day, 8 and 45 days after HIT compared to HIT only treatment. Apoptosis was also increased after SAHA only and HIT only 24 h, 8 and 45 days after irradiation but not as much as treatment with HIT and SAHA. Combination of HIT and SAHA lead to a significant (*) induction of necrosis on day 45. SAHA only and HIT only treatment resulted in a significantly higher rate of necrosis from day 24 on but not at earlier time points compared to the control groups

    Article Snippet: SAHA was obtained from Alexis Biochemicals (Lörrach, Germany), DMSO from Carl Roth Biochemicals (Karlsruhe, Germany).

    Techniques: Irradiation

    Expression of p53 and p21 WAF1/CIP1 is impaired in osteosarcoma xenografts after treatment with HIT and SAHA. Expression of p53 and p21 WAF1/CIP1 was analyzed after treatment with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. p53 expression was increased in all SAHA treated tumors compared to the vehicle treated control and only irradiated tumors on day 23 and 45. Increased p21 Waf1/Cip1 expression was detected in tumors after SAHA only and SAHA and HIT treatment compared to the controls and the HIT treated tumors on day 8

    Journal: Radiation Oncology (London, England)

    Article Title: Histone deacetylase inhibition sensitizes osteosarcoma to heavy ion radiotherapy

    doi: 10.1186/s13014-015-0455-z

    Figure Lengend Snippet: Expression of p53 and p21 WAF1/CIP1 is impaired in osteosarcoma xenografts after treatment with HIT and SAHA. Expression of p53 and p21 WAF1/CIP1 was analyzed after treatment with DMSO (control), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or HIT and SAHA. p53 expression was increased in all SAHA treated tumors compared to the vehicle treated control and only irradiated tumors on day 23 and 45. Increased p21 Waf1/Cip1 expression was detected in tumors after SAHA only and SAHA and HIT treatment compared to the controls and the HIT treated tumors on day 8

    Article Snippet: SAHA was obtained from Alexis Biochemicals (Lörrach, Germany), DMSO from Carl Roth Biochemicals (Karlsruhe, Germany).

    Techniques: Expressing, Irradiation

    Combination of HIT and SAHA induces an increased local control. Tumor growth of osteosarcoma xenografts was determined after treatment with DMSO (controls), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or SAHA plus HIT. Local control was defined as tumor growth > 1000 m 3 and calculated according to the method of Kaplan and Meier. The combination of HIT and SAHA led to a significant local control compared to SAHA only and HIT only starting day

    Journal: Radiation Oncology (London, England)

    Article Title: Histone deacetylase inhibition sensitizes osteosarcoma to heavy ion radiotherapy

    doi: 10.1186/s13014-015-0455-z

    Figure Lengend Snippet: Combination of HIT and SAHA induces an increased local control. Tumor growth of osteosarcoma xenografts was determined after treatment with DMSO (controls), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or SAHA plus HIT. Local control was defined as tumor growth > 1000 m 3 and calculated according to the method of Kaplan and Meier. The combination of HIT and SAHA led to a significant local control compared to SAHA only and HIT only starting day

    Article Snippet: SAHA was obtained from Alexis Biochemicals (Lörrach, Germany), DMSO from Carl Roth Biochemicals (Karlsruhe, Germany).

    Techniques: Irradiation

    The combination of HIT and SAHA results in a significant tumor growth delay compared to treatment with HIT or SAHA only. Osteosarcoma xenografts were treated with DMSO (controls), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or SAHA plus HIT and tumor growth was determined until day 45 after HIT. Comparing HIT as mono-treatment to SAHA only, HIT seemed to be superior from day 10 on after treatment start reaching significance at day 23. The combination of HIT and SAHA yielded a significant (*) tumor growth retardation compared to SAHA only and HIT only starting day 20 and day 25 respectively

    Journal: Radiation Oncology (London, England)

    Article Title: Histone deacetylase inhibition sensitizes osteosarcoma to heavy ion radiotherapy

    doi: 10.1186/s13014-015-0455-z

    Figure Lengend Snippet: The combination of HIT and SAHA results in a significant tumor growth delay compared to treatment with HIT or SAHA only. Osteosarcoma xenografts were treated with DMSO (controls), suberoylanilide hydroxamic acid (SAHA), irradiation (HIT) or SAHA plus HIT and tumor growth was determined until day 45 after HIT. Comparing HIT as mono-treatment to SAHA only, HIT seemed to be superior from day 10 on after treatment start reaching significance at day 23. The combination of HIT and SAHA yielded a significant (*) tumor growth retardation compared to SAHA only and HIT only starting day 20 and day 25 respectively

    Article Snippet: SAHA was obtained from Alexis Biochemicals (Lörrach, Germany), DMSO from Carl Roth Biochemicals (Karlsruhe, Germany).

    Techniques: Irradiation

    Gimatecan Inhibited HCC Cell Growth in Vitro . Different HCC cell lines were cultured and incubated with various concentrations of gimatecan at 37 ºC for 72 hours. The percentages of viable cells were measured at the end the incubation period. 100% refers to the number of cells after 72 hours of incubation in the presence of the vehicle control (0.1% DMSO).

    Journal: Asian Pacific Journal of Cancer Prevention : APJCP

    Article Title: In Vitro and In Vivo Anticancer Activity of Gimatecan against Hepatocellular Carcinoma

    doi: 10.22034/APJCP.2016.17.11.4853

    Figure Lengend Snippet: Gimatecan Inhibited HCC Cell Growth in Vitro . Different HCC cell lines were cultured and incubated with various concentrations of gimatecan at 37 ºC for 72 hours. The percentages of viable cells were measured at the end the incubation period. 100% refers to the number of cells after 72 hours of incubation in the presence of the vehicle control (0.1% DMSO).

    Article Snippet: Stock solutions of gimatecan were dissolved in 100% DMSO (Amresco, USA) and stored in sterilized brown glass bottles at -20ºC.

    Techniques: In Vitro, Cell Culture, Incubation

    In vivo Anti-Tumor Activity of Gimatecan in HCC Xenograft Models. Four types of HCC cell lines, Huh-1, Hep G2, HCCLM3, and PLCPRF5 were inoculated subcutaneously on the right side of the mouse back, respectively. In each tumor-bearing model mice were divided into four groups (n = 10 for each group) and orally administered with 0.8 mg/kg (×), 0.4 mg/kg (◆), and 0.1 mg/kg (●) gimatecan every four days for a total of four times, respectively. The control mice were administered with 10 µl 10%DMSO/g (●). Tumor volume was measured and expressed in mm 3

    Journal: Asian Pacific Journal of Cancer Prevention : APJCP

    Article Title: In Vitro and In Vivo Anticancer Activity of Gimatecan against Hepatocellular Carcinoma

    doi: 10.22034/APJCP.2016.17.11.4853

    Figure Lengend Snippet: In vivo Anti-Tumor Activity of Gimatecan in HCC Xenograft Models. Four types of HCC cell lines, Huh-1, Hep G2, HCCLM3, and PLCPRF5 were inoculated subcutaneously on the right side of the mouse back, respectively. In each tumor-bearing model mice were divided into four groups (n = 10 for each group) and orally administered with 0.8 mg/kg (×), 0.4 mg/kg (◆), and 0.1 mg/kg (●) gimatecan every four days for a total of four times, respectively. The control mice were administered with 10 µl 10%DMSO/g (●). Tumor volume was measured and expressed in mm 3

    Article Snippet: Stock solutions of gimatecan were dissolved in 100% DMSO (Amresco, USA) and stored in sterilized brown glass bottles at -20ºC.

    Techniques: In Vivo, Activity Assay, Mouse Assay

    Gimatecan Administration Did Not Severely Affect Body Weights. Four types of HCC cell lines, Huh-1, Hep G2, HCCLM3, and PLCPRF5 were inoculated subcutaneously on the right side of the mouse back, respectively. In each tumor-bearing model mice were divided into four groups (n = 10 for each group) and orally administered with 0.8 mg/kg (×), 0.4 mg/kg (◆), and 0.1 mg/kg (●) gimatecan every four days for a total of four times, respectively. The control mice were administered with 10 µl 10%DMSO/g (●). Mouse body weights were measured twice weekly and at study termination.

    Journal: Asian Pacific Journal of Cancer Prevention : APJCP

    Article Title: In Vitro and In Vivo Anticancer Activity of Gimatecan against Hepatocellular Carcinoma

    doi: 10.22034/APJCP.2016.17.11.4853

    Figure Lengend Snippet: Gimatecan Administration Did Not Severely Affect Body Weights. Four types of HCC cell lines, Huh-1, Hep G2, HCCLM3, and PLCPRF5 were inoculated subcutaneously on the right side of the mouse back, respectively. In each tumor-bearing model mice were divided into four groups (n = 10 for each group) and orally administered with 0.8 mg/kg (×), 0.4 mg/kg (◆), and 0.1 mg/kg (●) gimatecan every four days for a total of four times, respectively. The control mice were administered with 10 µl 10%DMSO/g (●). Mouse body weights were measured twice weekly and at study termination.

    Article Snippet: Stock solutions of gimatecan were dissolved in 100% DMSO (Amresco, USA) and stored in sterilized brown glass bottles at -20ºC.

    Techniques: Mouse Assay

    Analysis of RelA-dependent signaling in RelA-sfGFP-S4D-KI cells. a Quantitative RT-PCR for the expression of TNF-α-induced genes. Parental or RelA-sfGFP-S4D-KI cells were pretreated with DMSO or pomalidomide (Po) for 24 h. Then, the cells were stimulated with 20 ng/ml TNF-α for 1 h, and the expression of IκBα or IL-6 was measured by quantitative RT-PCR. The mRNA expression in untreated parental HeLa cells was set to 1.0. b , c Pomalidomide causes TNF-α-induced cell death. Parental and RelA-sfGFP-S4D-KI cells pretreated with DMSO or Po for 12 h were stimulated with 20 ng/ml TNF-α for 12 h, and the viability was measured by MTS assay ( b ) or trypan blue staining ( c ). d Immunoblot analysis of pomalidomide-dependent TNF-α-induced cell death. Parental and RelA-sfGFP-S4D-KI cells pretreated with pomalidomide for 12 h were stimulated with 50 ng/ml TNF-α for the indicated times, and effectors of cell death were analyzed by immunoblot. e , f zVAD-FMK treatment rescued TNF-α-induced cell death in pomalidomide-treated KI cells. Parental and RelA-sfGFP-S4D-KI cells pretreated with 10 µM Po for 12 h were then treated with DMSO or 10 µM zVAD-FMK. After 2 h of zVAD-FMK treatment, the cells were stimulated with 50 ng/ml TNF-α for 12 h and the viability was measured by trypan blue staining ( e ), or effectors of apoptosis were analyzed by immunoblot ( f ). Error bars in a – c and e represent the mean ± SD ( n = 3), and P values were calculated by one-way ANOVA with Tukey’s post-hoc tests (NS not significant; * P

    Journal: Communications Biology

    Article Title: An IMiD-induced SALL4 degron system for selective degradation of target proteins

    doi: 10.1038/s42003-020-01240-5

    Figure Lengend Snippet: Analysis of RelA-dependent signaling in RelA-sfGFP-S4D-KI cells. a Quantitative RT-PCR for the expression of TNF-α-induced genes. Parental or RelA-sfGFP-S4D-KI cells were pretreated with DMSO or pomalidomide (Po) for 24 h. Then, the cells were stimulated with 20 ng/ml TNF-α for 1 h, and the expression of IκBα or IL-6 was measured by quantitative RT-PCR. The mRNA expression in untreated parental HeLa cells was set to 1.0. b , c Pomalidomide causes TNF-α-induced cell death. Parental and RelA-sfGFP-S4D-KI cells pretreated with DMSO or Po for 12 h were stimulated with 20 ng/ml TNF-α for 12 h, and the viability was measured by MTS assay ( b ) or trypan blue staining ( c ). d Immunoblot analysis of pomalidomide-dependent TNF-α-induced cell death. Parental and RelA-sfGFP-S4D-KI cells pretreated with pomalidomide for 12 h were stimulated with 50 ng/ml TNF-α for the indicated times, and effectors of cell death were analyzed by immunoblot. e , f zVAD-FMK treatment rescued TNF-α-induced cell death in pomalidomide-treated KI cells. Parental and RelA-sfGFP-S4D-KI cells pretreated with 10 µM Po for 12 h were then treated with DMSO or 10 µM zVAD-FMK. After 2 h of zVAD-FMK treatment, the cells were stimulated with 50 ng/ml TNF-α for 12 h and the viability was measured by trypan blue staining ( e ), or effectors of apoptosis were analyzed by immunoblot ( f ). Error bars in a – c and e represent the mean ± SD ( n = 3), and P values were calculated by one-way ANOVA with Tukey’s post-hoc tests (NS not significant; * P

    Article Snippet: Reagents Thalidomide (Tokyo Chemical Industry), pomalidomide (Sigma–Aldrich), lenalidomide (FujiFilm Wako), 5-hydroxythalidomide (5-HT, synthesized as previously reported ), MG132 (Peptide Institute), MLN4924 (Chemscene), cycloheximide (Merck Millipore), and zVAD-FMK (Peptide Institute) were dissolved in DMSO (FujiFilm Wako) at 2–100 mM and stored at −20 °C as stock solutions.

    Techniques: Quantitative RT-PCR, Expressing, MTS Assay, Staining

    Construction of the IMiD-induced SALL4 degron (S4D) tag. a Amino acid sequence of IMiD-induced SALL4 degron. b Schematic diagram of the ORFs of the expression vector encoding Venus-SALL4 degron. c Immunoblot analysis of Venus-SALL4 degron protein. CRBN −/− HEK293T cells expressing AGIA-Venus-m1, -m2 (S4D), or -m3 and FLAG-CRBN were treated with DMSO (DM), thalidomide (Th), lenalidomide (Le), or pomalidomide (Po) for 16 h. d Immunoblot analysis of AGIA-Venus-S4D protein in CRBN −/− HEK293T cells expressing AGIA-S4D-WT and FLAG-CRBN treated with DM or Th in the presence of DM, MG132, or MLN4924 for 9 h. e Immunoblot analysis of Venus-S4D-WT or -G416A. CRBN −/− HEK293T cells expressing AGIA-Venus-S4D-WT or -G416A and FLAG-CRBN were treated with DM, Th, Le, or Po for 16 h. f Immunoblot analysis of Venus-S4D in HEK293T cells. HEK293T cells expressing AGIA-Venus-S4D were treated with DM, Th, Le, or Po for 16 h. g Schematic diagram of the ORF in an expression vector encoding FLuc and FLuc-S4D. h Immunoblot analysis of FLuc and FLuc-S4D. HEK293T cells expressing FLuc-AGIA or FLuc-S4D-AGIA were treated with DM, Th, Le, or Po for 16 h. i, j FLuc luciferase activity in lysates of HEK293T cells expressing FLuc-AGIA or FLuc-S4D-AGIA treated with DM, Th, Le, or Po for 16 h. k FLuc luciferase activity in lysates of HEK293T cells expressing FLuc-S4D-AGIA and treated with DMSO or 10 µM pomalidomide for the indicated times. l FLuc luciferase activity in lysates of HEK293T cells expressing FLuc-S4D-AGIA and treated with DM or Po in the presence of DM or MLN4924 for 12 h. Error bars in i-l represent the mean ± SD ( n = 3).

    Journal: Communications Biology

    Article Title: An IMiD-induced SALL4 degron system for selective degradation of target proteins

    doi: 10.1038/s42003-020-01240-5

    Figure Lengend Snippet: Construction of the IMiD-induced SALL4 degron (S4D) tag. a Amino acid sequence of IMiD-induced SALL4 degron. b Schematic diagram of the ORFs of the expression vector encoding Venus-SALL4 degron. c Immunoblot analysis of Venus-SALL4 degron protein. CRBN −/− HEK293T cells expressing AGIA-Venus-m1, -m2 (S4D), or -m3 and FLAG-CRBN were treated with DMSO (DM), thalidomide (Th), lenalidomide (Le), or pomalidomide (Po) for 16 h. d Immunoblot analysis of AGIA-Venus-S4D protein in CRBN −/− HEK293T cells expressing AGIA-S4D-WT and FLAG-CRBN treated with DM or Th in the presence of DM, MG132, or MLN4924 for 9 h. e Immunoblot analysis of Venus-S4D-WT or -G416A. CRBN −/− HEK293T cells expressing AGIA-Venus-S4D-WT or -G416A and FLAG-CRBN were treated with DM, Th, Le, or Po for 16 h. f Immunoblot analysis of Venus-S4D in HEK293T cells. HEK293T cells expressing AGIA-Venus-S4D were treated with DM, Th, Le, or Po for 16 h. g Schematic diagram of the ORF in an expression vector encoding FLuc and FLuc-S4D. h Immunoblot analysis of FLuc and FLuc-S4D. HEK293T cells expressing FLuc-AGIA or FLuc-S4D-AGIA were treated with DM, Th, Le, or Po for 16 h. i, j FLuc luciferase activity in lysates of HEK293T cells expressing FLuc-AGIA or FLuc-S4D-AGIA treated with DM, Th, Le, or Po for 16 h. k FLuc luciferase activity in lysates of HEK293T cells expressing FLuc-S4D-AGIA and treated with DMSO or 10 µM pomalidomide for the indicated times. l FLuc luciferase activity in lysates of HEK293T cells expressing FLuc-S4D-AGIA and treated with DM or Po in the presence of DM or MLN4924 for 12 h. Error bars in i-l represent the mean ± SD ( n = 3).

    Article Snippet: Reagents Thalidomide (Tokyo Chemical Industry), pomalidomide (Sigma–Aldrich), lenalidomide (FujiFilm Wako), 5-hydroxythalidomide (5-HT, synthesized as previously reported ), MG132 (Peptide Institute), MLN4924 (Chemscene), cycloheximide (Merck Millipore), and zVAD-FMK (Peptide Institute) were dissolved in DMSO (FujiFilm Wako) at 2–100 mM and stored at −20 °C as stock solutions.

    Techniques: Sequencing, Expressing, Plasmid Preparation, Luciferase, Activity Assay

    IMiD treatment induced the degradation of proteins with various subcellular localizations. a Schematic diagram of the ORFs of the expression vectors encoding p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, and MAVS-S4D-AGIA. b Localization of transiently expressed proteins. Immunofluorescent staining of p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA in HeLa cells. Scale bars, 20 µm. c Immunoblot analysis of the dose-dependent degradation of proteins with various subcellular localizations. HeLa cells expressing p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA and FLAG-CRBN were treated with DMSO (DM) or pomalidomide (Po) for 16 h. d Immunoblot analysis of the indicated proteins in various subcellular localizations. CRBN −/− HEK293T cells expressing p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA and FLAG-CRBN were treated with DM or Po in the presence of DM or 2 µM MLN4924 for 9 h. e Immunoblot analysis of CRBN dependency on protein degradation of proteins with various subcellular localizations. Parental or CRBN −/− HEK293T cells expressing p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA were treated with DM or Po for 16 h. f Schematic diagram of the ORFs of the expression vectors encoding GM130-S4D-AGIA. g Localization of transiently expressed GM130. Immunofluorescent staining of GM130-S4D-AGIA in HeLa cells. Scale bars, 20 µm. h HeLa cells expressing GM130-S4D-AGIA were treated with DMSO (DM) or pomalidomide (Po) for 16 h. i Immunoblot analysis of CRBN dependency on protein degradation of GM130-S4D-AGIA. GM130-S4D-AGIA degradation was analyzed by same procedure in Fig. 2e. j , k Immunoblot analysis of N- or C-terminally tagged p53 ( j ) or DRD1 ( k ). HEK293T cells expressing S4D-p53-AGIA, p53-S4D-AGIA, S4D-DRD1-AGIA, or DRD1-S4D-AGIA were treated with DM, thalidomide (Th), or Po for 16 h.

    Journal: Communications Biology

    Article Title: An IMiD-induced SALL4 degron system for selective degradation of target proteins

    doi: 10.1038/s42003-020-01240-5

    Figure Lengend Snippet: IMiD treatment induced the degradation of proteins with various subcellular localizations. a Schematic diagram of the ORFs of the expression vectors encoding p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, and MAVS-S4D-AGIA. b Localization of transiently expressed proteins. Immunofluorescent staining of p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA in HeLa cells. Scale bars, 20 µm. c Immunoblot analysis of the dose-dependent degradation of proteins with various subcellular localizations. HeLa cells expressing p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA and FLAG-CRBN were treated with DMSO (DM) or pomalidomide (Po) for 16 h. d Immunoblot analysis of the indicated proteins in various subcellular localizations. CRBN −/− HEK293T cells expressing p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA and FLAG-CRBN were treated with DM or Po in the presence of DM or 2 µM MLN4924 for 9 h. e Immunoblot analysis of CRBN dependency on protein degradation of proteins with various subcellular localizations. Parental or CRBN −/− HEK293T cells expressing p53-S4D-AGIA, STING-S4D-AGIA, DRD1-S4D-AGIA, or MAVS-S4D-AGIA were treated with DM or Po for 16 h. f Schematic diagram of the ORFs of the expression vectors encoding GM130-S4D-AGIA. g Localization of transiently expressed GM130. Immunofluorescent staining of GM130-S4D-AGIA in HeLa cells. Scale bars, 20 µm. h HeLa cells expressing GM130-S4D-AGIA were treated with DMSO (DM) or pomalidomide (Po) for 16 h. i Immunoblot analysis of CRBN dependency on protein degradation of GM130-S4D-AGIA. GM130-S4D-AGIA degradation was analyzed by same procedure in Fig. 2e. j , k Immunoblot analysis of N- or C-terminally tagged p53 ( j ) or DRD1 ( k ). HEK293T cells expressing S4D-p53-AGIA, p53-S4D-AGIA, S4D-DRD1-AGIA, or DRD1-S4D-AGIA were treated with DM, thalidomide (Th), or Po for 16 h.

    Article Snippet: Reagents Thalidomide (Tokyo Chemical Industry), pomalidomide (Sigma–Aldrich), lenalidomide (FujiFilm Wako), 5-hydroxythalidomide (5-HT, synthesized as previously reported ), MG132 (Peptide Institute), MLN4924 (Chemscene), cycloheximide (Merck Millipore), and zVAD-FMK (Peptide Institute) were dissolved in DMSO (FujiFilm Wako) at 2–100 mM and stored at −20 °C as stock solutions.

    Techniques: Expressing, Staining

    Generation and evaluation of knock-in (KI) cells expressing endogenous S4D-tagged protein. a Schematic diagram of S4D tag insertion at the C-terminus by genome editing using the CRISPR/Cas9 system. b Flowchart of the generation of sfGFP-S4D-tagged KI clones in HeLa cells. c Immunoblot analysis of S4D-tagged endogenous RelA and IκBα in KI cells. Parental, heterozygous, or homozygous KI cells were treated with DMSO (DM), 10 µM thalidomide (Th), or 10 µM pomalidomide (Po) for 24 h. d Immunoblot analysis of RelA-sfGFP-S4D and IκBα-sfGFP-S4D in each KI cell treated with DM or Po in the presence of DM, MG132, or MLN4924 for 9 h. e Dose-dependent degradation of S4D-tagged endogenous RelA and IκBα. Each KI cell was treated with DM, Th, or Po at the indicated concentration for 24 h, and the lysates were analyzed by immunoblot. f Time-dependent degradation of S4D-tagged endogenous RelA and IκBα. Each KI cell was treated with Po for the indicated times, and the lysates were analyzed by immunoblot. g Analysis of the reversibility of protein degradation in the S4D system by immunoblot. After each KI cell was treated with DM or Po for 6 h, the culture medium was replaced, and cells were harvested after the indicated times.

    Journal: Communications Biology

    Article Title: An IMiD-induced SALL4 degron system for selective degradation of target proteins

    doi: 10.1038/s42003-020-01240-5

    Figure Lengend Snippet: Generation and evaluation of knock-in (KI) cells expressing endogenous S4D-tagged protein. a Schematic diagram of S4D tag insertion at the C-terminus by genome editing using the CRISPR/Cas9 system. b Flowchart of the generation of sfGFP-S4D-tagged KI clones in HeLa cells. c Immunoblot analysis of S4D-tagged endogenous RelA and IκBα in KI cells. Parental, heterozygous, or homozygous KI cells were treated with DMSO (DM), 10 µM thalidomide (Th), or 10 µM pomalidomide (Po) for 24 h. d Immunoblot analysis of RelA-sfGFP-S4D and IκBα-sfGFP-S4D in each KI cell treated with DM or Po in the presence of DM, MG132, or MLN4924 for 9 h. e Dose-dependent degradation of S4D-tagged endogenous RelA and IκBα. Each KI cell was treated with DM, Th, or Po at the indicated concentration for 24 h, and the lysates were analyzed by immunoblot. f Time-dependent degradation of S4D-tagged endogenous RelA and IκBα. Each KI cell was treated with Po for the indicated times, and the lysates were analyzed by immunoblot. g Analysis of the reversibility of protein degradation in the S4D system by immunoblot. After each KI cell was treated with DM or Po for 6 h, the culture medium was replaced, and cells were harvested after the indicated times.

    Article Snippet: Reagents Thalidomide (Tokyo Chemical Industry), pomalidomide (Sigma–Aldrich), lenalidomide (FujiFilm Wako), 5-hydroxythalidomide (5-HT, synthesized as previously reported ), MG132 (Peptide Institute), MLN4924 (Chemscene), cycloheximide (Merck Millipore), and zVAD-FMK (Peptide Institute) were dissolved in DMSO (FujiFilm Wako) at 2–100 mM and stored at −20 °C as stock solutions.

    Techniques: Knock-In, Expressing, CRISPR, Clone Assay, Concentration Assay

    The combination of LCRF‐0006 and bortezomib synergistically increases MM tumor response in vivo. A, Schematic representation of treatment regimen. C57BL/KaLwRij mice with established MM (day 14 post‐5TGM1 cell injection) were treated with LCRF‐0006 (100 mg/kg) and bortezomib (0.5 mg/kg) (LCRF‐0006 + bortezomib), LCRF‐0006 and DMSO vehicle (LCRF‐0006), 2‐HP‐b‐CD vehicle and bortezomib (bortezomib) or 2‐HP‐b‐CD and DMSO vehicles (vehicles) i.p., as per treatment regimen. B, Tumor burden was assessed at day 14, 21 and 28 by BLI. Graph depicts mean ± SEM. n = 5‐6 mice/treatment group. *** P

    Journal: FASEB BioAdvances

    Article Title: LCRF‐0006, a small molecule mimetic of the N‐cadherin antagonist peptide ADH‐1, synergistically increases multiple myeloma response to bortezomib, et al. LCRF‐0006, a small molecule mimetic of the N‐cadherin antagonist peptide ADH‐1, synergistically increases multiple myeloma response to bortezomib

    doi: 10.1096/fba.2019-00073

    Figure Lengend Snippet: The combination of LCRF‐0006 and bortezomib synergistically increases MM tumor response in vivo. A, Schematic representation of treatment regimen. C57BL/KaLwRij mice with established MM (day 14 post‐5TGM1 cell injection) were treated with LCRF‐0006 (100 mg/kg) and bortezomib (0.5 mg/kg) (LCRF‐0006 + bortezomib), LCRF‐0006 and DMSO vehicle (LCRF‐0006), 2‐HP‐b‐CD vehicle and bortezomib (bortezomib) or 2‐HP‐b‐CD and DMSO vehicles (vehicles) i.p., as per treatment regimen. B, Tumor burden was assessed at day 14, 21 and 28 by BLI. Graph depicts mean ± SEM. n = 5‐6 mice/treatment group. *** P

    Article Snippet: 2.2 Drugs For in vitro experiments, LCRF‐0006 (kindly provided by Crocus Laboratories) was solubilized in dimethyl sulfoxide (DMSO) (Merck).

    Techniques: In Vivo, Mouse Assay, Injection