Kinetin was purchased from Nacalai Tesque (Tokyo, Japan). (−)-epigallocatechin gallate (EGCG), daidzein, genistein, epoxomicin, and bortezomib were purchased from FUJIFILM Wako Pure Chemical (Osaka, Japan). δ-tocotrienol was produced by LKT Laboratories, Inc. (St. Paul, MN). Phosphatidylserine was produced by Larodan Fine Chemicals AB (Solna, Sweden). Digoxin was produced by Cayman Chemical (Ann Arbor, MI). Carfilzomib was purchased from Cell Signaling Technology. b-RECTAS was synthesized in-house from RECTAS. Compound stock solutions were prepared with DMSO, except for Phosphatidylserine, which was dissolved in chloroform:methanol solution (95:5 %volume).
Bortezomib
Manufactured by Fujifilm
Sourced in Japan
Bortezomib is a laboratory equipment product manufactured by Fujifilm. It is a proteasome inhibitor that can be used for research purposes in various scientific applications.
Automatically generated - may contain errors
Lab products found in correlation
Lamin b1, by Cell Signaling Technology (2 mentions)
Fetal bovine serum (fbs), by Biowest (2 mentions)
Chloroquine, by Merck Group (2 mentions)
Mg132, by Merck Group (2 mentions)
Daidzein, by Fujifilm (1 mentions)
Genistein, by Fujifilm (1 mentions)
Digoxin, by Cayman Chemical (1 mentions)
Phosphatidylserine, by Larodan (1 mentions)
Sodium valproate, by Fujifilm (1 mentions)
5 azacytidine, by Fujifilm (1 mentions)
Fluorouracil 5 fu, by Fujifilm (1 mentions)
Cisplatin cddp, by Fujifilm (1 mentions)
Multiskan fc microplate photometer, by Thermo Fisher Scientific (1 mentions)
Celltiter 96 aqueous one solution reagent, by Promega (1 mentions)
N acetyl l cysteine nac, by Fujifilm (1 mentions)
Phorbol 12 myristate 13 acetate, by Merck Group (1 mentions)
Dynasore, by Merck Group (1 mentions)
Leupeptin, by Peptide Institute (1 mentions)
Epoxomicin, by Peptide Institute (1 mentions)
Nocodazole, by Merck Group (1 mentions)
9 protocols using bortezomib
1
Compound Procurement for Biological Assays
2
Investigating Cellular Protein Regulation
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All cultures and chemicals were purchased from Nacalai Tesque (Kyoto, Japan) unless otherwise indicated. Fetal bovine serum (FBS) was purchased from Bio West (Nuaillé, France). Bortezomib (a protease inhibitor) was obtained from Wako Chemicals (Osaka, Japan). TP and TOS were purchased from Sigma Aldrich (St. Louis, MO, USA), and T3 was purchased from Tama Biochemicals (Tokyo, Japan). Antibodies for NFE2L1 (#8052), ubiquitin (#3936) and Lamin B1 (#13435) were purchased from Cell Signaling Technology (Beverly, MA, USA). DDI2 antibody (sc-514004) is purchased from Santa Cruz Biotechnology (Dallas, TX, USA).
3
MTS Assay for Cell Proliferation and Cytotoxicity
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MTS assay using CellTiter 96 Aqueous One Solution Reagent (Promega, Madison, WI, USA), containing a tetrazolium compound [(3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt; MTS] and an electron coupling reagent (phenazine ethosulfate; PES) was adopted as a colorimetric method for determining the number of viable cells in the cell proliferation or cytotoxicity assays. The absorbance at 490nm was recorded using a Multiskan FC Microplate Photometer (Thermo Scientific).
For the proliferation assay, BAP1 knocked-down GBC cells were seeded on 96-well plates and measured with the MTS assay on days 0, 1, 3, 5, and 7. The data were presented as relative increases of the average intensity compared to the control group.
For drug sensitivity, BAP1 knocked-down cells were incubated on 96-well plates for 3 days after administration of the drugs, gemcitabine (GEM; Wako, Osaka, Japan), fluorouracil (5-FU; Wako), cisplatin (CDDP; Wako), sodium valproate (Wako), 5-azacytidine (Wako), and bortezomib (Wako). Cell viability was measured by MTS assay and the obtained data were analyzed by calculating the area under the curve (AUC) of each group using GraphPad Prism (ver.7.0, GraphPad Software, San Diego, CA, USA).
For the proliferation assay, BAP1 knocked-down GBC cells were seeded on 96-well plates and measured with the MTS assay on days 0, 1, 3, 5, and 7. The data were presented as relative increases of the average intensity compared to the control group.
For drug sensitivity, BAP1 knocked-down cells were incubated on 96-well plates for 3 days after administration of the drugs, gemcitabine (GEM; Wako, Osaka, Japan), fluorouracil (5-FU; Wako), cisplatin (CDDP; Wako), sodium valproate (Wako), 5-azacytidine (Wako), and bortezomib (Wako). Cell viability was measured by MTS assay and the obtained data were analyzed by calculating the area under the curve (AUC) of each group using GraphPad Prism (ver.7.0, GraphPad Software, San Diego, CA, USA).
4
Bortezomib and NAC Synergistic Treatment
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Bortezomib (021–18,901) and N‐acetyl‐l‐cysteine (NAC) (017–05131) were purchased from FUJIFILM Wako Pure Chemicals. Phorbol 12‐Myristate 13‐Acetate (P8139) was purchased from Sigma‐Aldrich.
5
Leptospire Infection Cellular Mechanisms
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The cells were infected with leptospires at a multiplicity of infection (MOI) of 100 from the basolateral side containing supplement-free DMEM/F-12 Ham medium. In the experiments involving the use of inhibitors, the inhibitors were added 30 min before infection. The inhibitors used were 30 μM dynasore (Sigma-Aldrich), 100 nM bafilomycin A1 (Bioviotica, Liestal, Switzerland), 50 μM chloroquine (Sigma-Aldrich), 10 μM MG-132 (Sigma-Aldrich), 200 nM bortezomib (FUJIFILM Wako, Osaka, Japan), 20 μM benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (Z-VAD-FMK, a pan-caspase inhibitor) (Tocris Bioscience, Bristol, UK), 20 μM benzyloxycarbonyl-Asp(OMe)-Glu(OMe)-Val-Asp(OMe)-fluoromethylketone (Z-DEVD-FMK, a caspase-3-specific inhibitor) (Tocris Bioscience), or 50 μM MDL-28170 (calpain inhibitor III) (Tokyo Chemical Industry, Tokyo, Japan). Cells were then incubated at 37°C and 5% CO2 during infection and lysed for proteome analysis and immunoblotting, or fixed for proximity ligation assay (PLA), immunostaining, and electron microscopy analysis.
6
Antibody Purification and Inhibitor Compounds
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Anti-Tks5 and anti-MT1-MMP polyclonal antibodies were raised by immunizing rabbits with purified recombinant His-tagged human Tks5 (aa 817–1118) and MT1-MMP (aa 280–520), respectively, and then the antisera were affinity-purified. Commercially available antibodies are listed in Table S6 . Secondary antibodies labeled with horseradish peroxidase (HRP) and fluorochrome were purchased from Bio-Rad Laboratories and Life Technologies, respectively. The following inhibitors were purchased: latrunculin A (Sigma-Aldrich), nocodazole (Sigma-Aldrich), MG132 (Sigma-Aldrich), epoxomicin (Peptide Institute), bortezomib (FujiFilm Wako), bafilomycin A1 (Millipore), chloroquine (Sigma-Aldrich), leupeptin (Peptide Institute), and wortmannin (Sigma-Aldrich).
7
Anticancer Agents Affecting Cellular Pathways
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All cultures and chemicals were purchased from Nacalai Tesque (Kyoto, Japan), unless otherwise indicated. Fetal bovine serum (FBS) was purchased from Bio West (Nuaillé, France). Cisplatin, pemetrexed, and bortezomib (a protease inhibitor) were obtained from Wako Chemicals (Osaka, Japan). Niclosamide (a STAT3 inhibitor) and marizomib (a proteasome inhibitor) were purchased from Abcam (Cambridge, UK) and MedChem Express (Monmouth Junction, NJ, USA), respectively. T3 was purchased from Tama Biochemicals (Tokyo, Japan). All antibodies, except for NRF1 and Lamin B1, were obtained from Cell Signaling Technology, Inc. (Beverly, MA, USA). Antibodies for NRF1 and Lamin B1 were purchased from ATLAS Antibodies (Stockholm, Sweden) and ProteinTech (Tokyo, Japan), respectively.
8
Everolimus and Bortezomib Synergistic Therapy
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Everolimus and bortezomib, purchased from Wako Pure Chemical Industries Ltd. (Osaka, Japan), were used for the investigation.
9
Investigating Anti-Cancer Compound Effects
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3‐Bromopyruvate (3‐BrPA) (16490) was purchased from Sigma‐Aldrich. Bortezomib (021‐18901) was purchased from FUJIFILM Wako Pure Chemical. Recombinant human interleukin‐6 (IL‐6) (#8904SC) was purchased from Cell Signaling Technology. AZD8055 was purchased from ChemieTek.
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