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Venetoclax

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Venetoclax is a small-molecule inhibitor that selectively binds to the B-cell lymphoma 2 (BCL-2) protein. It is designed to induce apoptosis in cancer cells that are dependent on BCL-2 for survival.

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Lab products found in correlation

Navitoclax, by Active Motif (3 mentions) A1331852, by Active Motif (2 mentions) Celltiter glo, by Promega (2 mentions) Anti vinculin, by Merck Group (1 mentions) Anti gapdh, by Merck Group (1 mentions) Thiazolyl blue tetrazolium bromide, by Merck Group (1 mentions) Tetramethylrhodamine methyl ester tmrm, by Thermo Fisher Scientific (1 mentions) Hoechst 33588, by Merck Group (1 mentions) Wehi 539, by ChemScene (1 mentions) 2 deoxy d glucose, by Merck Group (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) Propidium iodide, by Merck Group (1 mentions) Fetal bovine serum (fbs), by Merck Group (1 mentions) Rpmi 1640 medium, by Merck Group (1 mentions) D glucose, by Merck Group (1 mentions) Sodium pyruvate, by Merck Group (1 mentions) Dmem medium, by Lonza (1 mentions) Decitabine, by Selleck Chemicals (1 mentions) Zebularine, by Selleck Chemicals (1 mentions) Azacitidine, by Selleck Chemicals (1 mentions)

10 protocols using venetoclax

1

In vitro Sensitivity and Synergy in KMT2A-rearranged Infant ALL

Cited 4 times
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All in vitro sensitivity and synergy experiments were performed on an extensively characterized panel of eight patient-derived KMT2A-rearranged infant ALL cell lines (PER cell lines) [13 (link), 14 (link)], two commercially available KMT2A-rearranged infant ALL cell lines (ALL-PO and KOPN-8), and cells from four KMT2A-rearranged infant ALL patient-derived xenografts (MLL-5, MLL-7, MLL-14, and LR-iALL2) [15 (link), 16 (link)]. Assessment of in vitro sensitivity to azacitidine, decitabine, zebularine (Selleck Chemicals) and venetoclax (Active Biochem) and their synergy with each of the nine conventional chemotherapy agents used to treat infants with ALL, and synergy of each of the hypomethylating agents with venetoclax was performed as previously described [13 (link)]. In vitro drug combination studies were analyzed using SynergyFinder 2.0 [17 (link)]. Western blots were performed to detect DNMT1 levels following incubation with azacitidine and decitabine. Apoptosis and necrosis assays were performed using the RealTime-Glo™ Annexin V Apoptosis and Necrosis kit (Promega) according to the manufacturer’s instructions (Supplementary Methods).
Cheung L.C., Aya-Bonilla C., Cruickshank M.N., Chiu S.K., Kuek V., Anderson D., Chua G.A., Singh S., Oommen J., Ferrari E., Hughes A.M., Ford J., Kunold E., Hesselman M.C., Post F., Faulk K.E., Breese E.H., Guest E.M., Brown P.A., Loh M.L., Lock R.B., Kees U.R., Jafari R., Malinge S, & Kotecha R.S. (2022). Preclinical efficacy of azacitidine and venetoclax for infant KMT2A-rearranged acute lymphoblastic leukemia reveals a new therapeutic strategy. Leukemia, 37(1), 61-71.
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2

Immunoblotting of Cell Signaling Proteins

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Immunoblotting was performed with anti-GAPDH (Sigma-Aldrich, Munich, Germany), anti-vinculin (Sigma-Aldrich, Munich, Germany), anti-IP3R2 (Rbt02 [53 (link)]) and anti-Bim (Bioké, Leiden, The Netherlands).
BIRD-2 (RKKRRQRRRGGNVYTEIKCNSLLPLAAIVRV) (purity>85%) was purchased from LifeTein (South Plainfield, New Jersey, USA) and venetoclax from Active Biochem (Bonn, Germany).
Vervloessem T., Akl H., Tousseyn T., De Smedt H., Parys J.B, & Bultynck G. (2017). Reciprocal sensitivity of diffuse large B-cell lymphoma cells to Bcl-2 inhibitors BIRD-2 versus venetoclax. Oncotarget, 8(67), 111656-111671.
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3

Cytotoxicity Evaluation of Venetoclax and WEHI-539

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Fetal bovine serum, RPMI 1640 medium, Thiazolyl Blue Tetrazolium Bromide (MTT), dimethyl sulfoxide (DMSO), sodium pyruvate, D-glucose, 2-deoxy-D-glucose, Hoechst 33588 and propidium iodide (PI) came from Sigma-Aldrich (Dublin, Ireland). DMEM medium was purchased from Lonza (Analab Ltd, Lisburn, United Kindom). Tetramethylrhodamine methyl ester (TMRM) was from Invitrogen (Biosciences, Ireland). Venetoclax was purchased from Active Biochem (Maplewood, NJ, USA), WEHI-539 from ChemScene (South Brunswick, NJ, USA).
Lucantoni F., Düssmann H., Llorente-Folch I, & Prehn J.H. (2018). BCL2 and BCL(X)L selective inhibitors decrease mitochondrial ATP production in breast cancer cells and are synthetically lethal when combined with 2-deoxy-D-glucose. Oncotarget, 9(40), 26046-26063.
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4

Synthetic Lethal Validation via Combinatorial Screening

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Gene pairs associated with a synthetic lethal phenotype in the library screen were validated using combinatorial viability screening of sgRNA perturbations with 5 small molecule inhibitors: navitoclax (ABT-263, Active Biochem A-1001), A1331852 (Active Biochem A-6048); venetoclax (ABT-199, Active Biochem A-1231); WEHI539 (MedChem Express, HY-15607A); and the MCL1 inhibitor S63845 (a gift from Guo Wei, Golub lab). Meljuso cells were transduced in 12-well plates, as described above, with lentivirus containing a single sgRNA targeting one of the anti-apoptotic genes (BCL2L1, BCL2L2, MCL1) or a control sgRNA either targeting CD81 or containing a run of 6 thymidines. Two days after transduction, cells were selected using puromycin at 1 μg/mL for five days. After puromycin selection, 3,000 cells were seeded into 96-well plates. Across each row of the 96-well plate, a different small molecule was added at 11 log2-dilutions ranging from 1 μM to approximately 1 nM in duplicate for each of the cell lines. The last well in the row did not receive small molecule. After 3 days in the presence of the small molecule, viability of the cell population was assayed by CellTiterGlo (Promega) according to the manufacturer’s instructions.
Najm F.J., Strand C., Donovan K.F., Hegde M., Sanson K.R., Vaimberg E.W., Sullender M.E., Hartenian E., Kalani Z., Fusi N., Listgarten J., Younger S.T., Bernstein B.E., Root D.E, & Doench J.G. (2017). Orthologous CRISPR-Cas9 enzymes for Combinatorial Genetic Screens. Nature biotechnology, 36(2), 179-189.
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5

Simvastatin Activation and Small Molecule Modulation

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Simvastatin, atorvastatin calcium salt, rosuvastatin calcium salt, and fluvastatin sodium salt were obtained from Cayman Chemical Company. Simvastatin was activated as reported elsewhere (48 (link)). Venetoclax and navitoclax were obtained from Active Biochem. InSolution Q-VD-OPh was obtained from EMD Millipore. NVP-BEZ235 was obtained from LC Laboratories. Doxorubicin, vincristine, mevalonate, squalene, cholesterol, farnesyl pyrophosphate, and geranylgeranyl pyrophosphate were all obtained from Sigma-Aldrich.
Lee J.S., Roberts A., Juarez D., Vo T.T., Bhatt S., Herzog L.O., Mallya S., Bellin R.J., Agarwal S.K., Salem A.H., Xu T., Jia J., Li L., Hanna J.R., Davids M.S., Fleischman A.G., O’Brien S., Lam L.T., Leverson J.D., Letai A., Schatz J.H, & Fruman D.A. (2018). Statins enhance efficacy of venetoclax in blood cancers. Science translational medicine, 10(445), eaaq1240.
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6

Synthetic Lethal Validation via Combinatorial Screening

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Gene pairs associated with a synthetic lethal phenotype in the library screen were validated using combinatorial viability screening of sgRNA perturbations with 5 small molecule inhibitors: navitoclax (ABT-263, Active Biochem A-1001), A1331852 (Active Biochem A-6048); venetoclax (ABT-199, Active Biochem A-1231); WEHI539 (MedChem Express, HY-15607A); and the MCL1 inhibitor S63845 (a gift from Guo Wei, Golub lab). Meljuso cells were transduced in 12-well plates, as described above, with lentivirus containing a single sgRNA targeting one of the anti-apoptotic genes (BCL2L1, BCL2L2, MCL1) or a control sgRNA either targeting CD81 or containing a run of 6 thymidines. Two days after transduction, cells were selected using puromycin at 1 μg/mL for five days. After puromycin selection, 3,000 cells were seeded into 96-well plates. Across each row of the 96-well plate, a different small molecule was added at 11 log2-dilutions ranging from 1 μM to approximately 1 nM in duplicate for each of the cell lines. The last well in the row did not receive small molecule. After 3 days in the presence of the small molecule, viability of the cell population was assayed by CellTiterGlo (Promega) according to the manufacturer’s instructions.
Najm F.J., Strand C., Donovan K.F., Hegde M., Sanson K.R., Vaimberg E.W., Sullender M.E., Hartenian E., Kalani Z., Fusi N., Listgarten J., Younger S.T., Bernstein B.E., Root D.E, & Doench J.G. (2017). Orthologous CRISPR-Cas9 enzymes for Combinatorial Genetic Screens. Nature biotechnology, 36(2), 179-189.
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7

Venetoclax-Resistant Leukemia Cell Culture

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Riva WT and VR were grown in RPMI (Gibco/Invitrogen, Merelbeke, Belgium) supplemented with 2 mM glutamax, 10% heat-inactivated fetal bovine serum, 100 IU/mL penicillin and 100 µg/mL streptomycin (100× Pen/Strep, Gibco/Invitrogen, Merelbeke, Belgium) at 37 °C and 5% CO2. Resistance to Venetoclax was achieved by culturing Riva cells in increasing concentrations of Venetoclax over more than three months. This selection started with low nanomolar concentrations of Venetoclax (1–10 nM) and concentrations were stepwise increased up to 300 nM when more than 30% of cells survived the selection. Survival of cells was monitored using AnnexinV-FITC staining and flow cytometry twice a week. Venetoclax was purchased from Active Biochem (Kowloon, Hong Kong). Cells were seeded 24 h before being used in experiments at a density of 250,000/mL. BIRD-2 peptide (sequence: RKKRRQRRRGGNVYTEIKCNSLLPLAAIVRV) was purchased from LifeTein (South Plainfield, NJ, USA) with a purity of >85%. BDA-366 was purchased from BioVision (Milpitas, CA, USA). Both S63845 and A-1155463 were purchased from Selleckchem (Munich, Germany).
Kerkhofs M., Vervloessem T., Stopa K.B., Smith V.M., Vogler M, & Bultynck G. (2020). DLBCL Cells with Acquired Resistance to Venetoclax Are Not Sensitized to BIRD-2 But Can Be Resensitized to Venetoclax through Bcl-XL Inhibition. Biomolecules, 10(7), 1081.
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8

Novel Inhibitors in Cancer Research

Cited 1 time
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Venetoclax was purchased from Active Biochem (Bonn, Germany). A‐1331852 was purchased from Chemietek (Indianapolis, IN, USA). Ruxolitinib was purchased from Selleckchem (Houston, TX, USA). AS1517499 was purchased from Selleckchem. CW15337 was obtained from Prof. Dr. Simon Mackay (University of Strathclyde, Glasgow, UK) and previously described [15 (link)].
Haselager M.V., Thijssen R., Bax D., Both D., De Boer F., Mackay S., Dubois J., Mellink C., Kater A.P, & Eldering E. (2023). JAK–STAT signalling shapes the NF‐κB response in CLL towards venetoclax sensitivity or resistance via Bcl‐XL. Molecular Oncology, 17(6), 1112-1128.
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9

Peptide Synthesis and Compound Acquisition

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All peptides were custom synthesized by Mimotopes Pty Ltd (Australia). S55746 was purchased from ProbeChem (Cat#PC-63502). Venetoclax was purchased from ActiveBiochem (#A-1231). All other chemicals, unless specified, were obtained from Sigma-Aldrich (Australia). Primer sequences are detailed in Supplementary Table 3.
Birkinshaw R.W., Gong J.N., Luo C.S., Lio D., White C.A., Anderson M.A., Blombery P., Lessene G., Majewski I.J., Thijssen R., Roberts A.W., Huang D.C., Colman P.M, & Czabotar P.E. (2019). Structures of BCL-2 in complex with venetoclax reveal the molecular basis of resistance mutations. Nature Communications, 10, 2385.
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10

Eµ-Myc Lymphoma Cell Viability Assays

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Cell viability assays were performed on Eµ-Myc lymphoma cell lines to assess sensitivity to BH3 mimetic drug treatment. Cells were plated into 96-well flat-bottom plates at a density of 3 × 104 cells per well and treated for 24 h with the MCL-1 inhibitor S63845 (Active Biochem, #A-6044) or the BCL-2 inhibitor venetoclax (Active Biochem, #A-1231) at the indicated concentrations. Cell viability was determined by staining with 1 µg/mL propidium iodide (PI, Sigma–Aldrich, #P4864) followed by flow cytometric analysis using the LSR-II Analyzer (BD Biosciences). Flow cytometry data were analysed using FlowJo v10 software (BD Biosciences).
Deng Y., Diepstraten S.T., Potts M.A., Giner G., Trezise S., Ng A.P., Healey G., Kane S.R., Cooray A., Behrens K., Heidersbach A., Kueh A.J., Pal M., Wilcox S., Tai L., Alexander W.S., Visvader J.E., Nutt S.L., Strasser A., Haley B., Zhao Q., Kelly G.L, & Herold M.J. (2022). Generation of a CRISPR activation mouse that enables modelling of aggressive lymphoma and interrogation of venetoclax resistance. Nature Communications, 13, 4739.
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