Medicine with non-small cell lung cancer resisting effect and use thereof
A non-small cell lung cancer, drug technology, applied in the direction of antineoplastic drugs, drug combinations, medical preparations containing active ingredients, etc., can solve the problems of drug resistance side effects, death, treatment can not continue, etc., to achieve strong anti-proliferation activity Effect
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Embodiment 1
[0019] Embodiment 1: NCI-H1975 (EGFR L858R / T790M ), HCC827 (EGFR E746-A750del ) and A549 (EGFR WT ) cell screening to optimize gefitinib Gef and febuxostat FBS composition
[0020] Take the cells in the logarithmic growth phase and inoculate 3×10 4 Cells / well were placed on a 96-well plate. After 6 hours of growth, the supernatant was discarded by centrifugation, and then administered according to the following groups: Tumor cells were divided into no drug group and drug-added group, and the drug-added group consisted of Gef and FBS single drugs Group, Gef and FBS combined drug groups with different molar ratios, set 4-6 duplicate wells in each group, culture for 24 hours, discard the supernatant, add 100 μl of MTT (tetrazolium salt) serum-free culture medium containing 0.5 mg / ml for culture After 4 hours, add 100 μl of DMSO (dimethyl sulfoxide), place on a micro-oscillator to vibrate for 10 minutes, and then place on a microplate reader to detect the OD value at 570 nm. R...
Embodiment 2
[0026] Embodiment 2: Expression of EGFR and Western blot detection
[0027] Immune Western blot antibodies were purchased from Abcam's C-Caspase-3 and anti-EGFR (ab52894) (Abcam). After the three kinds of cells were treated with gefitinib Gef or febuxostat FBS for 48 hours, 5×10 6 Cells were lysed with 200 μl of RIPA (50 mM Tris pH 8.0, 150 mM NaCl, 0.1% SDS, 0.5% sodiumdeoxycholate, 1% NP-40) and added protease inhibitors on ice for 30 minutes, then centrifuged at 12,000 rpm for 10 minutes, and the supernatant was collected for BCA method To measure protein concentration, add 2×SDS loading buffer and cook at 100°C for 10 minutes. The total protein (100 μg) was loaded, followed by 8-15% SDS-PAGE gel electrophoresis according to the molecular weight of the protein, and then transferred to a PVDF membrane (GE healthcare), incubated with corresponding antibodies, and the chemiluminescent enzyme substrate HRP Substrate ( Millipore Corporation), applied LAS-4000 imaging system im...
Embodiment 3
[0028] Example 3: FBS drug inhibits tumor growth in non-small cell lung cancer tumor-bearing mice
[0029] In order to test the efficacy of FBS on non-small cell lung cancer in vivo, we established a mouse tumor model of non-small cell lung cancer in nude mice using NCI-H1975 cells. Including control group (Control) (physiological saline); FBS (300mg / kg / d, intragastric administration) and Gef (25mg / kg / d, intragastric administration) groups and the Gef / FBS combination group with the same dose as single use, each group 10 nude mice, after subcutaneous inoculation of tumors, when the subcutaneous tumor volume is greater than 100mm 3 At that time, the mice were randomly divided into four groups, and intragastric administration was started after the 7th day, once a day, and the body weight change and tumor volume were weighed the next day, and the suspension was diluted with normal saline. The major diameter and minor diameter of the tumor were measured with a vernier caliper ever...
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