Osimertinib-resistant human non-small cell lung cancer cell strain H1975/OR and application thereof

A non-small cell lung cancer, lung cancer cell technology, applied in the field of human non-small cell lung cancer cell line H1975/OR, can solve the problem of unclear drug resistance mechanism

Active Publication Date: 2020-10-20
SUN YAT SEN UNIV CANCER CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Among patients with acquired resistance mutations of the first generation EGFR-TKI, about 50% of patients will have T970M point mutations. Patients with T790M mutations can choose the third generation EGFR-TKI osimertinib. After 10 months of sigtinib, drug resistance will still appear again, and the mechanism of drug resistance has not been clarified in the prior art

Method used

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  • Osimertinib-resistant human non-small cell lung cancer cell strain H1975/OR and application thereof
  • Osimertinib-resistant human non-small cell lung cancer cell strain H1975/OR and application thereof
  • Osimertinib-resistant human non-small cell lung cancer cell strain H1975/OR and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1 Preparation of osimertinib-resistant human non-small cell lung cancer cell line H1975 / OR

[0022] The preparation method comprises the following steps:

[0023] (1) Human EGFR-positive non-small cell lung cancer cell line H1975 (hereinafter referred to as human non-small cell lung cancer cell line H1975) was purchased from American Type Culture Collection (ATCC). Place the cell H1975 strain in RPMI1640 culture medium containing 25 mmol / L 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) and 10% fetal bovine serum (FBS) (hereinafter referred to as H1975 medium) at 37°C, 5% CO 2 , cultured in an incubator with saturated humidity, digested and passaged with 0.25% trypsin and ethylenediaminetetraacetic acid (0.25% Trypsin-EDTA);

[0024] (2) Take about 5×10 5 A H1975 cell was placed in a culture flask, and after 24 hours of attachment, the old culture medium was aspirated, and the newly prepared H1975 medium containing 0.1 μmol / L osimertinib was added,...

Embodiment 2

[0026] Example 2 Cryopreservation of osimertinib-resistant human non-small cell lung cancer cell line H1975 / OR of the present invention

[0027] The drug-resistant cells obtained in Example 1 were placed in the cryopreservation solution of the cryopreservation tube, and then the cryopreservation tube was placed in liquid nitrogen for preservation. The cryopreservation medium consists of 90% H1975 medium and 10% dimethyl sulfoxide (DMSO). The cryopreservation process takes the steps of placing the cryopreservation tube in a nalgene programmed cooling box, overnight at -80°C, and finally putting it into liquid nitrogen.

Embodiment 3

[0028] Example 3 Recovery of osimertinib-resistant human non-small cell lung cancer cell line H1975 / OR

[0029]Take out the cryopreservation tube containing the cells from the liquid nitrogen, immediately put it in a 37-40°C water bath and shake it gently, so that the frozen storage will thaw within 1 minute, wipe the outer wall of the cryopreservation tube with an alcohol cotton ball and put it into the ultra-clean tower. Put the thawed cell suspension into a 15ml sterile centrifuge tube, add 5ml of H1975 medium, centrifuge at 1200 rpm for 5 minutes, discard the supernatant, add 5ml of H1975 medium containing 10μmol / L osimertinib, and Pipette and mix to form a mixed solution, suck the mixed solution and put it into a 25cm 2 In the cell culture bottle, unscrew the bottle cap and put it in a carbon dioxide incubator at 37°C, 5% CO 2 1. Cultivate under the condition of saturated humidity, replace the medium once every 3-4 days, digest when the cell density reaches 80-90%, and ...

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Abstract

The invention discloses an osimertinib-resistant human non-small cell lung cancer cell strain which is named as osimertinib-resistant human lung cancer cell line H1975 / OR, and has a preservation number of CCTCC NO:C2019300. According to the cell strain, EGFR exon21: c.T2573G: p.L858R mutation and exon20: c.C2369T: p.T790M mutation which are sensitive to osimertinib are reserved, RB1 gene deletionoccurs after drug resistance is acquired, and a small cell lung cancer phenotype is shown. The osimertinib-resistant human non-small cell lung cancer cell strain H1975 / OR disclosed by the invention can be used for researching the morphological and biological characteristics of osimertinib-resistant human non-small cell lung cancer cells, studying a tumor drug resistance mechanism, analyzing the sensitivity of the anti-tumor drug, screening and evaluating the anti-tumor drug, developing a tumor drug resistance reversal drug and studying a more effective tumor treatment method, and can be used for discussion of a small cell lung cancer transformation mechanism after EGFR-TKIs treatment and research of related signal channels, has high scientific research and production application values, and is expected to produce good scientific research, economic and social benefits.

Description

technical field [0001] The invention relates to the field of tumor biology, in particular to an osimertinib-resistant human non-small cell lung cancer cell line H1975 / OR and an application thereof. Background technique [0002] Lung cancer is a malignant tumor with the highest morbidity and mortality in the world. Lung cancer is divided into small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC), of which about 85% are non-small cell lung cancer. The main treatment methods for non-small cell lung cancer include molecular targeted therapy, immunotherapy, chemotherapy and radiotherapy. Among them, molecular targeted therapy has the advantages of high specificity and mild adverse reactions. [0003] In recent years, epidermal growth factor tyrosine kinase inhibitor (EGFR-TKI) has become the first choice for the treatment of advanced non-small cell lung cancer with EGFR gene mutation. EGFR-TKI can compete with the ATP site in the EGFR tyrosine kinase region, blo...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12Q1/02C12Q1/18C12R1/91
CPCC12N5/0693C12N5/0688G01N33/5011G01N33/5044C12N2503/02G01N2500/10
Inventor 张力潘慧袁泽占建华黄岩
Owner SUN YAT SEN UNIV CANCER CENT
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