Application of combination of sorafenib and oleanolic acid in preparation of medicine for treating liver cancer
An oleanolic acid, a technology for treating liver cancer, applied in the field of medicine, can solve the problems of SOR toxicity, easy to produce drug resistance, etc.
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Embodiment 1
[0041] Example 1 BEL-7402 cell culture
[0042] BEL-7402 was cultured with RPMI 1640 medium containing 10% fetal bovine serum (FBS), 100×L-glutamine (L-Glu), and 100×HEPES. Routinely take 400,000 BEL-7402 cells in the logarithmic growth phase and inoculate them in a 60mm plate, 3ml system, and pass them every 2 days. When passaging, first remove the old medium, and then use 1:1 diluted trypsin digestion solution (Trypsin-EDTA 0.25% to HBSS volume ratio 1:1). After 1ml digestion for 2 minutes, discard the trypsin digestion solution and add 1ml fresh culture Base, gently blow down the adherent cells to mix evenly, passaging 1:4 to 1:5.
Embodiment 2
[0043] Example 2 Cell Proliferation Inhibition Experiment
[0044] In order to observe whether the combination of SOR and OA has a synergistic effect, the BEL-7402 cells prepared in Example 1 were used for different administration treatments, and the cell proliferation activity was detected by the CCK-8 method, and the proliferation of BEL-7402 cells was inhibited by the action of drugs In order to judge the efficacy of the combined application of the two drugs. The specific operation is as follows:
[0045] (1) Plating: Specify the total system of 200μl / well in a 96-well plate. The BEL-7402 cells in the logarithmic growth phase were seeded in a 96-well plate (1000 cells / well, 100 μl cell suspension), and incubated overnight at 37°C until the cells adhered.
[0046] (2) Dosing treatment: set the empty well group (only containing 100μl of medium), blank control group (BEL-7402 cells, medium), solvent control (BEL-7402 cells, medium, solvent, the solvent is the volume 1:1 ratio of D...
Embodiment 3
[0062] Example 3 Experiment on the influence of cell cycle
[0063] SOR and OA were used in combination to act on BEL-7402 cells, and flow cytometry was used for analysis to determine the effect of the combination of the two drugs on the cell cycle progression of BEL-7402.
[0064] (1) Preparation of cell samples to be tested
[0065] (1) Inoculate BEL-7402 cells in logarithmic growth phase in a 6-well plate, 200,000 per well, 2ml culture system, and culture at 37°C for 24 hours.
[0066] (2) SOR (2,4,10μM), OA (2,5,10μM) and SOR 4μM are used in combination with OA (2,5,10μM) respectively. The maximum amount of working solution is 1ml, and each is prepared (2×)10μM SOR and (2×)10μM OA working liquid were administered sequentially in gradients, and 2ml of the total system was supplemented with culture medium (same as Example 1) so that the final concentration of working liquid in each well after administration was set as above. Constant temperature treatment at 37°C for 24h.
[0067] (...
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