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Drug-resistance cell system for lung cancer and preparation method thereof

A technology for drug-resistant cells and lung cancer cells, which can be applied to tumors/cancer cells, biochemical equipment and methods, animal cells, etc., can solve problems such as drug resistance in patients with lung cancer, and achieve easy industrial preparation, volume increase, and invasion. Ability-enhancing effect

Inactive Publication Date: 2018-06-05
SHANGHAI CHEST HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Osimertinib (AZD9291) is a third-generation targeted drug with significant curative effect after EGFR-TKI resistance, and it can simultaneously treat EGFR gene mutations (including exon 18, 19, and 21 mutations) in non-small cell lung cancer and EGFR-TKI acquired drug resistance (T790M), prolonging the life of patients with advanced lung cancer, however, it is inevitable that lung cancer patients will still develop drug resistance after a period of time

Method used

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  • Drug-resistance cell system for lung cancer and preparation method thereof
  • Drug-resistance cell system for lung cancer and preparation method thereof
  • Drug-resistance cell system for lung cancer and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] After resuscitating the lung cancer cell line H1975 cells, use RPMI1640 culture medium containing 10% fetal bovine serum, and incubate at 37°C, 5% CO 2 cultured in an incubator.

[0044] Take the H1975 cell line in the exponential growth phase 1×10 6 Cells / ml were seeded in a 6-well plate, and an appropriate amount of RPMI1640 medium containing osimertinib (AZD9291) was added, placed at 37°C, 5% CO 2 cultured in a humidified incubator. The initial concentration of AZD9291 added was 93 nM.

[0045]Observe the condition of the cells after 72 hours of action, and when the cells return to normal growth, add the drug at the same concentration again until the cells survive and proliferate normally at this concentration, which is regarded as a culture cycle.

[0046] Continue to increase the drug concentration and carry out the next cycle. During the initial screening, increase the concentration difference by every 50nM. After 24 hours of treatment, replace the complete med...

Embodiment 2

[0051] After resuscitating the lung cancer cell line HCC827 cells, use RPMI1640 culture medium containing 10% fetal bovine serum, and incubate at 37°C, 5% CO 2 cultured in an incubator.

[0052] Take 1×10 HCC827 cell line in exponential growth phase 6 Cells / ml were seeded in a 6-well plate, and an appropriate amount of RPMI1640 medium containing osimertinib (AZD9291) was added, placed at 37°C, 5% CO 2 cultured in a humidified incubator. The initial concentration of AZD9291 added was 21 nM.

[0053] Observe the condition of the cells after 72 hours of action, and when the cells return to normal growth, add the drug at the same concentration again until the cells survive and proliferate normally at this concentration, which is regarded as a culture cycle.

[0054] Continue to increase the drug concentration and carry out the next cycle. During the initial screening, increase the concentration difference by every 50nM. After 24 hours of treatment, replace the complete medium w...

experiment example 1

[0059] The functional experiment of the drug-resistant cell line that embodiment 1 and 2 make

[0060] (1) Changes in cell morphology

[0061] The drug-resistant cells after successful screening have morphological changes compared with non-resistant cells, such as cells becoming larger and spindle-shaped. Non-drug-resistant cells and drug-resistant cells were observed and photographed using an inverted microscope. Such as figure 1 shown. figure 1 Among them, A is H1975 normal cells; B is H1975 / RS; C is HCC827; D is HCC827 / RS.

[0062] figure 1 The results showed that the volume of the drug-resistant cell group was larger than that of the non-drug-resistant cell group, and the H1975 / RS cells showed an obvious spindle shape. The above results confirmed that the cell strains with characteristics of drug-resistant cells were successfully obtained.

[0063] (2) Cell proliferation assay

[0064] Drug-resistant cells have the characteristics of long proliferation cycle and slo...

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Abstract

The invention relates to the field of drug-resistance cell systems, and particularly relates to a drug-resistance cell system and a preparation method thereof. The method comprises the following steps: inoculating the lung cancer cells to a culture medium containing a 21-93nM lung cancer targeted drug; culturing until cells can normally live and multiply under the same concentration; improving andculturing at 50nM concentration difference; improving and culturing at concentration difference of every 100nM when the cell multiplying inhibiting concentration is 4 times of IC50; improving and culturing at concentration difference of every 1 microns when the drug sieving concentration is 8 times of IC50; and obtaining the drug-resistance cell systems for the lung cancer when the lung cancer cells can live and multiply in a cell culture system of 10 microns osimertinib. According to the method, the drug-resistance cell system is prepared on the basis of gradually increasing the concentration of drugs in the culture medium; the foundation is provided for later simulation of clinical drug resistance, researching of drug resistance mechanism of lung cancer targeted drugs, and developing ofeffective drug resisting drugs.

Description

technical field [0001] The invention relates to the field of drug-resistant cell lines, in particular to a lung cancer drug-resistant cell line and a preparation method thereof. Background technique [0002] Lung cancer is one of the most common malignant tumors in the world. It can be divided into small cell lung cancer and non-small cell lung cancer. Among them, non-small cell lung cancer accounts for about 80% of all lung cancers. According to the National Cancer Center in 2017, lung cancer has become the cancer with the highest morbidity and mortality in China, and most patients are diagnosed at an advanced stage. The treatment of advanced lung cancer is mostly through chemotherapy, but due to the heterogeneity and variability of tumor cells, many tumor patients are not sensitive to chemotherapy drugs, or gradually develop tolerance to drugs during chemotherapy, resulting in tumor recurrence or death of the patient. The drug resistance of lung cancer tumors is mainly d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/09
CPCC12N5/0693C12N2500/30
Inventor 杨运海陈天翔
Owner SHANGHAI CHEST HOSPITAL
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