Molecular Weight(MW): 520.62
Purmorphamine, which directly binds and activates Smoothened, blocks BODIPY-cyclopamine binding to Smo with IC50 of ~ 1.5 μM in HEK293T cell and also is an inducer of osteoblast differentiation with EC50 of 1 μM.
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(C) mRNA levels of Nek2A in purmorphamine treated H4 cells. Total mRNA was extracted and quantified by real-time PCR. Error bars represent the standard deviation of three independent experiments. **P<0.01 compared with the control groups.
Int J Oncol, 2016, doi: 10.3892/ijo.2016.3819. . Purmorphamine purchased from Selleck.
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|Description||Purmorphamine, which directly binds and activates Smoothened, blocks BODIPY-cyclopamine binding to Smo with IC50 of ~ 1.5 μM in HEK293T cell and also is an inducer of osteoblast differentiation with EC50 of 1 μM.|
Purmorphamine activates the Hedgehog pathway by directly binds and activates Smoothened with IC50 of ~ 1.5 μM in compete with cyclopamine, a Smo antagonist.  Purmorphamine is a potent inducer of osteogenesis in multipotent C3H10T1/2 cells. The EC50 (based on ALP expression) for Purmorphamine is 1 μM in C3H10T1/2 cells. Purmorphamine (1 μM) and BMP-4 (100 ng/mL) together increase ALP activity more than 90-fold in 3T3-L1 cells.  In contrast to BMP-4, Purmorphamine induces osteogenesis by activating Hedgehog signaling in multipotent mesenchymal progenitor cells. 
|In vivo||Purmorphamine up-regulates ALP expression in human mesenchymal stem cell-based constructs on rats. |
Binding assay:Smo binding assays are conducted with BODIPY-cyclopamine and Smo-overexpressing cells as previously described4,5, using CMV promoter-based, SV40 origin-containing expression constructs for Smo-Myc3, the deletion mutant SmoCRD (deletion of amino acids 68 to 182), and SmoCT (deletion of amino acids 556 to 793). HEK 293T cells are grown on poly-D-lysine-treated glass coverslips in 12-well plates until 70% confluency and then transfected with the appropriate expression construct (0.5 g/well) using FuGene 6 according the manufacturer's protocols. Two days after transfection, the HEK 293T cells are incubated with DMEM containing 0.5% bovine calf serum, 5 nM BODIPY-cyclopamine, and varying concentrations of Purmorphamine (0, 1.5, or 5 M) (1 mL/well) for 1 h at 37 ℃. The Smo-overexpressing cells are then washed with 1 × PBS buffer (1 mL/well), mounted with DAPI-containing medium, and visualized using a Leica DM4500B fluorescence microscope. For binding assays using fixed cells, the Smo-overexpressing HEK 293T cells are fixed with 3% paraformaldehyde in 1 × PBS buffer for 10 min at room temperature (1 mL/well), treated with 1 × PBS containing 10 mM glycine and 0.2% sodium azide for 5 min (1 mL/well), washed with 1 × PBS buffer (1 mL/well), and treated with the Purmorphamine-containing media described above for 4 h at room temperature.
|In vitro||DMSO||4 mg/mL warmed (7.68 mM)|
|In vivo||2% DMSO+30% PEG 300+5% Tween 80+ddH2O||1mg/mL|
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