Xmd8 92

Manufactured by Santa Cruz Biotechnology

Sourced in United States

XMD8-92 is a small molecule inhibitor that targets the protein TANK-binding kinase 1 (TBK1). It is a laboratory research tool used in the study of TBK1 and its role in various cellular processes.

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4 protocols using xmd8 92

Molecular Markers of Cellular Signaling

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An Annexin V-FITC kit was obtained from BD Biosciences. Antibodies against ERK5, phospho-ERK5, ERK1/2, phospho-ERK1/2, phospho-checkpoint kinase 1 (Chk1), phospho-checkpoint kinase 2 (Chk2), phosphor-ATM kinase, phosphor-ATR protein, cleaved caspase-9, cleaved caspase-8, cleaved caspase-3, Cyclin B1, Cdc-2, and Cdc25C were purchased from Cell Signaling Technologies. Antibodies against p53, α-tubulin, β-actin, and cleaved poly (ADP-ribose) polymerase (PARP), and the ERK5 inhibitor XMD8-92 were purchased from Santa Cruz Biotechnology. Horseradish peroxidase (HRP)-conjugated goat anti-rabbit and anti-mouse IgG antibodies were also obtained from Santa Cruz Biotechnology. Anti-H2A histone family, member X (H2AX, Ser139) was provided by Millipore (Billerica, MA, USA). Alexa Fluor 488 goat anti-mouse IgG (H+L) secondary antibody was purchased from Invitrogen (Carlsbad, CA, USA). Anti-CD31 antibody was obtained from Abcam (Cambridge, MA, USA). Sources of other materials are noted accordingly in the study.

Jiang W., Jin G., Cai F., Chen X., Cao N., Zhang X., Liu J., Chen F., Wang F., Dong W., Zhuang H, & Hua Z.C. (2019). Extracellular signal-regulated kinase 5 increases radioresistance of lung cancer cells by enhancing the DNA damage response. Experimental & Molecular Medicine, 51(2), 1-20.

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Benzidine-induced molecular signaling

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Benzidine (4, 4′-diaminobiphenyl; ≥98.0%, RT), methanol as well as DMSO were purchased from Merck (USA). Benzidine was dissolved in DMSO, which was subsequently preserved at −20°C. The final concentration of DMSO administered in cells was under 1‰. Polyclonal antibodies against p-ERK1/2, p-p38, p-JNK, p-ERK5, p-c-Jun, p-c-Fos, and Fra-1 were purchased from Cell Signaling Technology (USA). Polyclonal antibodies against ZO-1, E-cadherin, Snail, N-cadherin, Vimentin, XMD8-92, U0126, as well as GAPDH were commercially obtained from Santa Cruz (USA). Primers for Snail, N-cadherin, Vimentin, ZO-1, E-cadherin as well as GAPDH were synthesized by Invitrogen (USA). Sources of unmentioned materials were described in the following.

Sun X., Zhang T., Deng Q., Zhou Q., Sun X., Li E., Yu D, & Zhong C. (2018). Benzidine Induces Epithelial–Mesenchymal Transition of Human Bladder Cancer Cells through Activation of ERK5 Pathway. Molecules and Cells, 41(3), 188-197.

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Molecular Targeting and Analysis

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Tamoxifen, safflower oil, DL-dithiothreitol (DTT), DMSO and fluorescein isothiocyanate (FITC)-Dextran (FD-4) were purchased from Sigma. EDTA was obtained from Invitrogen. MEK1/2 inhibitor PD0325901 was obtained from Stemgent. ERK5 inhibitor XMD8-92 was obtained from Santa Cruz Biotechnology. EGFR inhibitor erlotinib was obtained from Selleckchem. Recombinant mWnt3a was obtained from R&D Systems.

de Jong P.R., Taniguchi K., Harris A.R., Bertin S., Takahashi N., Duong J., Campos A.D., Powis G., Corr M., Karin M, & Raz E. (2016). ERK5 signalling rescues intestinal epithelial turnover and tumour cell proliferation upon ERK1/2 abrogation. Nature Communications, 7, 11551.

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Cell Proliferation Assay with Kinase Inhibitors

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PD98059, a special inhibitor of the ERK1/2, was from (Sigma Company), SP600125, the inhibitor of JNK, was from (Sigma Company), XMD8-92, an effective inhibitor of the ERK5, from (Santa Cruz Biotechnology). Treated with different concentration of ERK1/2 inhibitor, JNK inhibitor, ERK5 inhibitor, cell proliferation ability of each group were assessed using MTT assay. Briefly, the exponential growth phase, were trypinized, centrifuged at 400 × g for 5 min at room temperature, resuspend in complete medium (10% FBS) and were counted. The cells were then seeded into five 96-well plates (1×10³ cells/well), with five parallel wells for each cell group. The medium was replaced with 120 µl MTT solution, containing 100 µl medium and 20 µl MTT (Beijing Solarbio Science & Technology Co., Ltd.), at different time-points (24, 48, 72 h). After 4 h, the medium was aspirated and 150 µl DMSO was added to each well, then the plate was agitated for 45 sec at 27°C. The optical density value at a wavelength of 490 nm was determined using a Multiskan FC Microplate photometer (Thermo Fisher Scientific, Waltham, MA, USA).

Wang Z., Wang W., Xu S., Wang S., Tu Y., Xiong Y., Mei J, & Wang C. (2016). The role of MAPK signaling pathway in the Her-2-positive meningiomas. Oncology Reports, 36(2), 685-695.

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