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Gastrointestinal stromal tumor drug-resistant cell model as well as construction method and application thereof

A technology for gastrointestinal stromal tumors and drug-resistant cells, which is applied to gastrointestinal cells, tumor/cancer cells, cells modified by introducing foreign genetic material, etc., can solve the problem of sunitinib-resistant cell lines that have not been reported And other issues

Active Publication Date: 2021-01-12
SHANGHAI PUTUO DISTRICT LIQUN HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there are no reports of sunitinib-resistant cell lines

Method used

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  • Gastrointestinal stromal tumor drug-resistant cell model as well as construction method and application thereof
  • Gastrointestinal stromal tumor drug-resistant cell model as well as construction method and application thereof
  • Gastrointestinal stromal tumor drug-resistant cell model as well as construction method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] 1. Materials and methods

[0028] 1.1 GIST882 cell culture: The cell line was imported from ATCC / Chinese Academy of Sciences / Union Hospital, and the cell STR identification has been carried out. The cell culture medium was complete DMEM medium containing 10% fetal bovine serum. Place the cells at 37°C, 5% CO 2 And cultivated in the incubator with saturated humidity.

[0029] 1.2 Induction of drug-resistant cell lines: The concentration gradient method was used to establish the Sunitinib-resistant cell line GIST882-Rs of gastrointestinal stromal tumors. Take GIST882 under the logarithmic growth phase, add DMEM culture solution containing sunitinib, and continue to act at a low concentration (1 / 2IC50 concentration) for 24-48h, the sensitive cells begin to apoptosis, discard the culture solution, and wash with PBS Wash the buffer 3 times, change the fresh culture medium, continue to culture the drug-resistant cells in the culture medium (without drug) until entering the...

Embodiment 2

[0043] We expanded the gastrointestinal stromal tumor Sunitinib-resistant cell line GIST882-Rs and stored it in a -80°C refrigerator. After 6 and 12 months, we recovered and detected the IC50 of Sunitinib7 to evaluate the stability of the cell line. Results After 6 months of storage, the IC50 was 129.61±1.86 μmol / L, and after 6 months of storage, the IC50 was 126.31±1.10 μmol / L. The drug resistance did not fluctuate greatly, indicating good stability.

Embodiment 3

[0045] Our research group constructed 9 Sunitinib7-resistant GIST882 cell lines before and after using the concentration gradient method. The Sunitinib-resistant cell line GIST882-Rs of the gastrointestinal stromal tumor in Example 1 was one of the lines constructed in the third batch. The drug resistance was the strongest, and the IC50 of the remaining 8 Sunitinib7-resistant cells were 69.5, 51.1, 97.2, 80.6, 66.5, 73.9, and 102.5 μmol / L when they were established.

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Abstract

The invention relates to a gastrointestinal stromal tumor drug-resistant cell model as well as a construction method and application thereof. In-vitro induction is performed on gastrointestinal stromal tumor cell strains GIST882 by an intermittent concentration gradient multiplication method, a drug-resistant cell line with Sunitinib IC50 of 127.5 + / -2.15 [mu] mol / L and a drug resistance index of5.30 is established, the cell line changes in cell morphology, ultrastructure and cell cycles, Kit gene sequencing shows that 17 exon R804L mutation exists, INPP4B gene sequencing shows that 18 exon V466I mutation exists, and besides, IRS2, SOX9, FLCN, WISP3, SOX10 and MYCN also have mutation difference. The invention lays a foundation for further researching the drug resistance mechanism of the Sunitinib and screening new drugs for treating gastrointestinal stromal tumors.

Description

technical field [0001] The invention relates to the technical field of cell models, in particular to a drug-resistant cell model of gastrointestinal stromal tumor and its construction method and application. Background technique [0002] Gastrointestinal Stromal Tumors (GIST), as a common foliar-derived tumor, is insensitive to radiotherapy and chemotherapy. Sunitinib is the main targeted therapy drug for the treatment of advanced gastrointestinal stromal tumors. . Sunitinib is a multi-target inhibitor that can effectively inhibit receptors encoded by KIT, VEGFR, PDGFRa, FLT3 and RET proto-oncogenes. With the increasingly obvious problem of drug resistance in the clinical use of Sunitinib, it is necessary to conduct in-depth research and analysis on the specific drug resistance mechanism. It is difficult to obtain secondary samples from drug-resistant patients, and the amount of tissue is difficult to meet the needs of our experiments. Therefore, drug-resistant cell lines ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12Q1/02C12R1/91
CPCC12N5/0693C12N5/0679G01N33/5011C12N2510/00G01N2500/10
Inventor 王翠华于学涛王婷琦赵胜伟姚新明战海峰王伟冯丽波钱小峰
Owner SHANGHAI PUTUO DISTRICT LIQUN HOSPITAL
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