A kind of aurora A protein inhibitor and its preparation method and pharmaceutical application
A kind of use and medicine technology, applied in the field of AuroraA protein inhibitor and preparation thereof
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Embodiment 1
[0051] Embodiment 1: synthetic Py-Im-Ht (synthetic route sees figure 1 )
[0052] 1. Synthesis of Precursor 1
[0053] Add hydrazine resin (0.61mmol / g, 400mg, 0.244mmol) and dichloromethane (3ml) into a 50mL solid phase reactor, and swell the resin for 20min. Extract dichloromethane, add 20% piperidine / DMF solution (3ml) to the resin, blow air for 5min, remove solvent, then add 20% piperidine / DMF solution (3ml), blow air for 5min, remove solvent, The resin was washed with DMF (4x 3 mL). Boc-Py-OH (234mg, 0.976mmol, 4eq.) and triphosgene (BTC, 95mg, 0.322mmol, 0.33eq) were dissolved in 2mL of anhydrous THF, and collidine (collidine, 284μL, 2.928mmol, 12eq.), the reaction immediately produces a large amount of white precipitate, after adding the reaction for 3min, then add 2mL DIEA / DMF solution (5%, v / v), the white precipitate completely disappears, transfer the reaction solution to the In the phenylhydrazine resin of the protective group, N2 was bubbled and mixed, and the ...
Embodiment 2
[0069] Embodiment 2: Py-Im-Ht inhibits Aurora A protein expression experiment in tumor cells (HeLa and A549)
[0070]Take Hela cells and A549 cells in the logarithmic growth phase, inoculate 1 mL per well (50,000 cells) in a 12-well plate, and culture them in a 37-degree, 5% carbon dioxide incubator for 24 hours; The original culture medium was added to DMEM high-glucose medium containing different concentrations of Compound 2, so that the drug concentration in each well was 0 μM, 10 μM, 20 μM, and 30 μM; the cells were put back into the incubator and incubated for 72 hours; Digest with 0.25% trypsin at room temperature for 1 min, resuspend the cells in fresh DMEM medium, centrifuge at 3000 rpm for 3 minutes, collect the cells, and wash twice with ice-cold PBS. Remove PBS, centrifuge at 5000rpm for 3 minutes, add RIPA lysate, incubate on ice for 2 hours, centrifuge at 13000rpm at 4 degrees for 15min, collect the supernatant, measure the protein concentration by BCA method, add...
Embodiment 3
[0071] Example 3: Experiment of Py-Im-Ht inhibiting tumor cell proliferation and cell cycle
[0072] Take tumor cells (HeLa, A549 or MDA-MB-231) in the logarithmic growth phase, inoculate 1 mL per well (containing 100,000 cells) in a 6-well plate or 0.1 mL per well (containing 5,000 cells) In a 96-well plate, cultured in a 37°C, 5% carbon dioxide cell incubator for 24 hours; aspirated the original medium, and added it to DMEM high-glucose medium containing different concentrations of Py-Im-Ht; put the cells back into Incubate in an incubator for 48 hours to 72 hours, use a microscope to count the final cell number and the number of cells in each division period or use the CCK-8 kit to measure cell viability, the results are as follows Figure 4 shown. It can be seen that with the prolongation of drug action time, the number of Hela cells and A549 cells decreased significantly ( Figure 4 A&B), indicating that the drug can effectively inhibit the proliferation of tumor cells....
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