Bufadienolide compound and application thereof to preparation of medicine for resisting gastric cancer
A technology of bufadienolide and compound is applied in the field of anti-gastric cancer drugs, which can solve the problems of unreported anti-gastric cancer activity of bufadienolide compounds.
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Embodiment 1
[0014] Example 1: Establishment of a 3D cell spheroid screening model for gastric cancer
[0015] Sand toad toxin, sand toad toxin-3-pimeloyl arginine ester, far cinobufin toxin-3-adipyl arginine ester, far cinobufin toxin, deacetylated cinobufin toxin, Chinese Bufafen-3-suberoyl arginine ester, bufafen-3-suberoyl arginine ester, 3-dehydrobufafolin and sea bufo toxin were sourced from Shanghai Yuanye Biotechnology Co., Ltd. .
[0016] Docetaxel and epirubicin hydrochloride were purchased from Dalian Meilun Biotechnology Co., Ltd.; gastric cancer cell line HGC-27 was purchased from Shanghai Cell Bank, Chinese Academy of Sciences; ULA-96-well plates were purchased from Corning; inverted microscopes were purchased from OLYMPUS.
[0017] The HGC-27 cells in the logarithmic growth phase were seeded in ULA-96 well plates at a density of 20000 cells / well (20K cells / well), 10000 cells / well, 5000 cells / well, 2500 cells / well, and 1250 cells / well. Cells / well, 625 cells / well, 313 cells / ...
Embodiment 2
[0018] Example 2: Inhibitory effect of coarsely screened bufadienolactone compounds on the growth of 3D cell spheres
[0019] Docetaxel and epirubicin hydrochloride, typical drugs for the clinical treatment of gastric cancer, were selected as positive control drugs, and the inhibitory effect of 9 typical bufadiene lactone compounds on the growth of HGC-27 3D cell spheres was roughly screened. Among them, 9 typical bufadiene lactone compounds (as a coarse screening drug) are bufabufagin, bufabufalin-3-pimeloyl arginate, and bufabufalin-3-adipyl Arginine Esters, Cinobufagin, Deacetylated Cinobufagin, Cinobufagin-3-Suberoyl Arginate, Cinobufagin-3-Suberoyl Arginate, 3 - Dehydrobufalin and sea bufotoxin. Cells at the optimal density (313 cells / well) were inoculated on ULA-96 well plates, and the inoculation volume of the medium (RPMI-1640 containing 20% fetal bovine serum) was 200 μL / well. After culturing for 4 days and growing to 3D cell spheres with optimal particle size, 10...
Embodiment 3
[0020] Example 3: The dose relationship of deacetylated cinobufagin inhibiting the growth of 3D cell spheres
[0021] Different doses of the compound with the strongest inhibitory ability, 5-deacetylcinobufagin, were applied to the 3D cell spheres. Inoculate the cells with optimal density (313 cells / well) in ULA-96 well plate, culture medium (RPMI-1640 containing 20% fetal calf serum) with an inoculation volume of 200 μL / well for 4 days, and grow to the optimal particle size After 3D cell spheres, replace 100 μL of fresh medium in each well, add compound 5 to 16 wells with a volume of 10 μL, and the concentration gradient of the compound is 40 μM, 20 μM, 10 μM, 5 μM, 2.5 μM, 1.25 μM, 0.625 μM and 0.3125 μM, each concentration was replicated twice. The blank control group was cell spheres treated with medium containing 0.2% DMSO (RPMI-1640 containing 20% fetal bovine serum), and two wells were used as the blank control group. Use an inverted microscope to record the size ...
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