MiR-517a-3p related to cisplatin resistance of tumor cells and application of miR-517a-3p

A technology of mir-517a-3p and tumor cells, applied in the field of biomedicine, can solve problems affecting tumor treatment effects, tumor drug resistance, etc.

Active Publication Date: 2021-02-12
河北仁博科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, like chemotherapy, tumor drug resistance also occurs during targeted therapy, which has become a major obstacle affecting the efficacy of tumor therapy

Method used

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  • MiR-517a-3p related to cisplatin resistance of tumor cells and application of miR-517a-3p
  • MiR-517a-3p related to cisplatin resistance of tumor cells and application of miR-517a-3p
  • MiR-517a-3p related to cisplatin resistance of tumor cells and application of miR-517a-3p

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 The cultivation of SK-OV-3 cells and the establishment of SK-OV-3 / DDP drug-resistant cell lines

[0027] 1. Culture of SK-OV-3 cells

[0028] SK-OV-3 (ovarian cancer cells) were cultured in DMEM / F12 medium containing 10% FBS at 37°C, 5% CO 2 cultured in an incubator.

[0029] 2. Establishment of SK-OV-3 / DDP drug-resistant cell lines

[0030] Screen drug-resistant cell lines by continuously increasing the concentration of drugs.

[0031] First, the IC50 of cisplatin (Yuan Ye Bio, B24462) in SK-OV-3 cells (ovarian cancer cells) is 500nM. The DDP concentration of SK-OV-3 cells in the logarithmic growth phase starts from 500nM, and cultures for 3 days. The cells are replaced with fresh culture medium to remove dead cells and continue to culture without adding drugs. After 1 to 2 weeks of culture, the surviving cells gradually grow into cell clusters. After trypsinization and passaging, add drugs and continue to culture for 1 to 2 weeks, and the surviving cells...

Embodiment 2

[0032] Example 2 Detection of differential expression of miR-517a-3p in SK-OV-3 wild cells and SK-OV-3 / DDP cells

[0033] The expression difference of miR-517a-3p in SK-OV-3 wild cells and drug-resistant cells SK-OV-3 / DDP was detected by fluorescent quantitative PCR.

[0034] 1. miRNA extraction

[0035] The control group SK-OV-3 wild cells and SK-OV-3 cisplatin-resistant cells were collected respectively. Using the miRNA extraction kit from Tiangen, add 1ml of lysate to every 50mg of cell sample, shake and mix for 30 seconds. After standing at room temperature for 5 minutes, centrifuge at 12,000rpm for 10min, take the supernatant, add 200μl chloroform, and shake vigorously for 15 seconds. After standing at room temperature for 5min, centrifuge at 12,000rpm for 15min. Water phase, RNA is mainly in the water phase, transfer the water phase to a new tube, slowly add 1 / 3 volume of absolute ethanol to mix well, transfer to the adsorption column together, place at room temperatur...

Embodiment 3

[0045] Example 3 Effect of interfering with miR-517a-3p on cell drug resistance

[0046] By inhibiting the function of the miRNA, observe whether it can reduce the drug resistance of tumor cells.

[0047] Cells in the logarithmic growth phase were taken for cell transfection, and miR-517a-3p inhibitor and miR-NC inhibitor (Guangzhou Ruibo Biotechnology Co., Ltd.) were transfected into SK-OV-3 according to the instructions of the transfection reagent lipofectamine 3000 / DDP-resistant cells.

[0048] (1) CCK8 method was used to detect the half inhibitory concentration (IC50) of cells to drugs before and after miR-517a-3p interference.

[0049] After 24 hours of transfection, the cells were digested with 0.25% trypsin and prepared into a single cell suspension, and the cells were seeded in 96-well culture plates, 6x10 3 cells / well. After the cells adhered to the wall, DDP with a final concentration of 5 nmol / L, 4.5 nmol / L, 4 nmol / L, 3.5 nmol / L, 3 nmol / L, 2.5 nmol / L, and 2 nmol...

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Abstract

The invention discloses miR-517a-3p related to cisplatin resistance of tumor cells and an application of the miR-517a-3p. The invention also discloses a kit for detecting the drug resistance degree oftumor cells to cisplatin. According to the miR-517a-3p, by establishing an SK-OV-3/DDP drug-resistant cell strain, it is found that miR-517a-3p can be used as a marker of drug resistance of tumor cells; the miR-517a-3p plays a certain role in the drug resistance of the SK-OV-3/DDP cells to the DDP, and the drug resistance of the SK-OV-3/DDP cells to the DDP can be reversed by down-regulating themiR-517a-3p; in addition, the proliferation speed of ovarian cancer SK-OV-3/DDP cells can be inhibited, and meanwhile, the cloning formation capability of the ovarian cancer SK-OV-3/DDP cells is alsoinhibited. The miR-517a-3p is advantaged in that a scientific basis is provided for searching a new molecular treatment target, guiding clinical medication and more effectively resisting cisplatin resistance.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to miR-517a-3p related to tumor cell cisplatin resistance and application thereof. Background technique [0002] Malignant tumor (cancer) is a common disease that seriously endangers human life and health. According to the data released by the World Health Organization, there are about 12.7 million new cancer cases in the world every year, and about 7.6 million people die of cancer. It is estimated that New cases and deaths will continue to rise, and by 2030, more than 13.1 million people will die from cancer each year. Conventional treatments for cancer include surgery, chemotherapy, and radiation therapy. Chemotherapy, also known as drug therapy, has always played an important role in cancer treatment, but its therapeutic effect is affected by its dose-dependent toxicity. At present, the effect of drug therapy has entered a plateau. Molecular targeted therapy refers to usi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886A61K45/06A61K33/243A61K31/7088A61P35/00
CPCC12Q1/6886A61K45/06A61K33/243A61K31/7088A61P35/00C12Q2600/106C12Q2600/158C12Q2600/178A61K2300/00
Inventor 崔健
Owner 河北仁博科技有限公司
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